• Of mice and men: the host response to influenza virus infection.

      Kollmus, Heike; Pilzner, Carolin; Leist, Sarah R; Heise, Mark; Geffers, Robert; Schughart, Klaus; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-08-01)
      Influenza virus (IV) infections represent a very serious public health problem. At present, no established biomarkers exist to support diagnosis for respiratory viral infections and more importantly for severe IV disease. Studies in animal models are extremely important to understand the biological, genetic, and environmental factors that contribute to severe IV disease and to validate biomarker candidates from human studies. However, mouse human cross-species comparisons are often compromised by the fact that animal studies concentrate on the infected lungs, whereas in humans almost all studies use peripheral blood from patients. In addition, human studies do not consider genetic background as variable although human populations are genetically very diverse. Therefore, in this study, we performed a cross-species gene expression study of the peripheral blood from human patients and from the highly genetically diverse Collaborative Cross (CC) mouse population after IV infection. Our results demonstrate that changes of gene expression in individual genes are highly similar in mice and humans. The top-regulated genes in humans were also differentially regulated in mice. We conclude that the mouse is a highly valuable in vivo model system to validate and to discover gene candidates which can be used as biomarkers in humans. Furthermore, mouse studies allow confirmation of findings in humans in a well-controlled experimental system adding enormous value to the understanding of expression and function of human candidate genes.
    • OLYMPUS: An automated hybrid clustering method in time series gene expression. Case study: Host response after Influenza A (H1N1) infection

      Dimitrakopoulou, Konstantina; Vrahatis, Aristidis G.; Wilk, Esther; Tsakalidis, Athanasios K.; Bezerianos, Anastasios (2013-10-31)
    • The phosphatidylserine receptor has essential functions during embryogenesis but not in apoptotic cell removal

      Böse, Jens; Gruber, Achim D; Helming, Laura; Schiebe, Stefanie; Wegener, Ivonne; Hafner, Martin; Beales, Marianne; Köntgen, Frank; Lengeling, Andreas (BioMed Central, 2004)
      Background Phagocytosis of apoptotic cells is fundamental to animal development, immune function and cellular homeostasis. The phosphatidylserine receptor (Ptdsr) on phagocytes has been implicated in the recognition and engulfment of apoptotic cells and in anti-inflammatory signaling. To determine the biological function of the phosphatidylserine receptor in vivo, we inactivated the Ptdsr gene in the mouse. Results Ablation of Ptdsr function in mice causes perinatal lethality, growth retardation and a delay in terminal differentiation of the kidney, intestine, liver and lungs during embryogenesis. Moreover, eye development can be severely disturbed, ranging from defects in retinal differentiation to complete unilateral or bilateral absence of eyes. Ptdsr -/- mice with anophthalmia develop novel lesions, with induction of ectopic retinal-pigmented epithelium in nasal cavities. A comprehensive investigation of apoptotic cell clearance in vivo and in vitro demonstrated that engulfment of apoptotic cells was normal in Ptdsr knockout mice, but Ptdsr-deficient macrophages were impaired in pro- and anti-inflammatory cytokine signaling after stimulation with apoptotic cells or with lipopolysaccharide. Conclusion Ptdsr is essential for the development and differentiation of multiple organs during embryogenesis but not for apoptotic cell removal. Ptdsr may thus have a novel, unexpected developmental function as an important differentiation-promoting gene. Moreover, Ptdsr is not required for apoptotic cell clearance by macrophages but seems to be necessary for the regulation of macrophage cytokine responses. These results clearly contradict the current view that the phosphatidylserine receptor primarily functions in apoptotic cell clearance.
    • PLAU inferred from a correlation network is critical for suppressor function of regulatory T cells.

      He, Feng; Chen, Hairong; Probst-Kepper, Michael; Geffers, Robert; Eifes, Serge; Del Sol, Antonio; Schughart, Klaus; Zeng, An-Ping; Balling, Rudi; 1] Department of Infection Genetics, Helmholtz Centre for Infection Research (HZI), University of Veterinary Medicine Hannover, Braunschweig, Germany [2] Luxembourg Centre for Systems Biomedicine (LCSB), University of Luxembourg, Esch-sur-Alzette, Luxembourg. (2012-11-20)
      Human FOXP3(+)CD25(+)CD4(+) regulatory T cells (Tregs) are essential to the maintenance of immune homeostasis. Several genes are known to be important for murine Tregs, but for human Tregs the genes and underlying molecular networks controlling the suppressor function still largely remain unclear. Here, we describe a strategy to identify the key genes directly from an undirected correlation network which we reconstruct from a very high time-resolution (HTR) transcriptome during the activation of human Tregs/CD4(+) T-effector cells. We show that a predicted top-ranked new key gene PLAU (the plasminogen activator urokinase) is important for the suppressor function of both human and murine Tregs. Further analysis unveils that PLAU is particularly important for memory Tregs and that PLAU mediates Treg suppressor function via STAT5 and ERK signaling pathways. Our study demonstrates the potential for identifying novel key genes for complex dynamic biological processes using a network strategy based on HTR data, and reveals a critical role for PLAU in Treg suppressor function.
    • Protection from Severe Influenza Virus Infections in Mice Carrying the Mx1 Influenza Virus Resistance Gene Strongly Depends on Genetic Background.

      Shin, Dai-Lun; Hatesuer, Bastian; Bergmann, Silke; Nedelko, Tatiana; Schughart, Klaus; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2015-10)
      Influenza virus infections represent a serious threat to human health. Both extrinsic and intrinsic factors determine the severity of influenza. The MX dynamin-like GTPase 1 (Mx1) gene has been shown to confer strong resistance to influenza A virus infections in mice. Most laboratory mouse strains, including C57BL/6J, carry nonsense or deletion mutations in Mx1 and thus a nonfunctional allele, whereas wild-derived mouse strains carry a wild-type Mx1 allele. Congenic C57BL/6J (B6-Mx1(r/r)) mice expressing a wild-type allele from the A2G mouse strain are highly resistant to influenza A virus infections, to both mono- and polybasic subtypes. Furthermore, in genetic mapping studies, Mx1 was identified as the major locus of resistance to influenza virus infections. Here, we investigated whether the Mx1 protective function is influenced by the genetic background. For this, we generated a congenic mouse strain carrying the A2G wild-type Mx1 resistance allele on a DBA/2J background (D2-Mx1(r/r)). Most remarkably, congenic D2-Mx1(r/r) mice expressing a functional Mx1 wild-type allele are still highly susceptible to H1N1 virus. However, pretreatment of D2-Mx1(r/r) mice with alpha interferon protected them from lethal infections. Our results showed, for the first time, that the presence of an Mx1 wild-type allele from A2G as such does not fully protect mice from lethal influenza A virus infections. These observations are also highly relevant for susceptibility to influenza virus infections in humans.
    • The proteolytic activation of A (H3N2) Influenza virus hemagglutinin is facilitated by different type II transmembrane serine proteases.

      Kühn, Nora; Bergmann, Silke; Kasnitz, Nadine; Lambertz, Ruth L O; Keppner, Anna; van den Brand, Judith M A; Pöhlmann, Stefan; Weiß, Siegfried; Hummler, Edith; Hatesuer, Bastian; Schughart, Klaus; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2016-02-17)
      Cleavage of influenza virus hemagglutinin (HA) by host cell proteases is necessary for viral activation and infectivity. In humans and mice, members of the type II transmembrane protease family (TTSP), e.g. TMPRSS2, TMPRSS4 and TMPRSS11d (HAT), have been shown to cleave influenza virus HA for viral activation and infectivity in vitro. Recently, we reported that inactivation of a single HA-activating protease gene, Tmprss2, in knock-out mice inhibits spread of H1N1 influenza viruses. However, after infection of Tmprss2 knock-out mice with H3N2 only a slight increase was observed in survival and mice still lost body weight. In this study, we investigated an additional trypsin-like protease, TMPRSS4. Both TMPRSS2 and TMPRSS4 are expressed in the same cell types of the mouse lung. Deletion of Tmprss4 alone in knock-out mice does not protect them from body weight loss and death upon infection with H3N2 influenza virus. In contrast, Tmprss2(-/-)Tmprss4(-/-) double knock-out mice showed a remarkably reduced virus spread and lung pathology in addition to reduced body weight loss and mortality. Thus, our results identified TMPRSS4 as a second host cell protease that, in addition to TMPRSS2, is able to activate the HA of H3N2 influenza virus HA in vivo.
    • QTLminer: identifying genes regulating quantitative traits

      Alberts, Rudi; Schughart, Klaus (2010-10-15)
      Abstract Background Quantitative trait locus (QTL) mapping identifies genomic regions that likely contain genes regulating a quantitative trait. However, QTL regions may encompass tens to hundreds of genes. To find the most promising candidate genes that regulate the trait, the biologist typically collects information from multiple resources about the genes in the QTL interval. This process is very laborious and time consuming. Results QTLminer is a bioinformatics tool that automatically performs QTL region analysis. It is available in GeneNetwork and it integrates information such as gene annotation, gene expression and sequence polymorphisms for all the genes within a given genomic interval. Conclusions QTLminer substantially speeds up discovery of the most promising candidate genes within a QTL region.
    • QTLminer: identifying genes regulating quantitative traits.

      Alberts, Rudi; Schughart, Klaus; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2010-10-15)
      Quantitative trait locus (QTL) mapping identifies genomic regions that likely contain genes regulating a quantitative trait. However, QTL regions may encompass tens to hundreds of genes. To find the most promising candidate genes that regulate the trait, the biologist typically collects information from multiple resources about the genes in the QTL interval. This process is very laborious and time consuming.
    • Quantitative determination of the diagnostic accuracy of the synovitis score and its components.

      Slansky, Elisabeth; Li, Jialiang; Häupl, Thomas; Morawietz, Lars; Krenn, Veit; Pessler, Frank; Medical Faculty 'Carl Gustav Carus', Technical University of Dresden, Germany. (2010-09)
      To assess the diagnostic accuracy of a three-component synovitis score and to determine the relative contribution of each of its components to its overall discriminatory power.
    • Respiratory Mucosal Proteome Quantification in Human Influenza Infections.

      Marion, Tony; Elbahesh, Husni; Thomas, Paul G; DeVincenzo, John P; Webby, Richard; Schughart, Klaus; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2016)
      Respiratory influenza virus infections represent a serious threat to human health. Underlying medical conditions and genetic make-up predispose some influenza patients to more severe forms of disease. To date, only a few studies have been performed in patients to correlate a selected group of cytokines and chemokines with influenza infection. Therefore, we evaluated the potential of a novel multiplex micro-proteomics technology, SOMAscan, to quantify proteins in the respiratory mucosa of influenza A and B infected individuals. The analysis included but was not limited to quantification of cytokines and chemokines detected in previous studies. SOMAscan quantified more than 1,000 secreted proteins in small nasal wash volumes from infected and healthy individuals. Our results illustrate the utility of micro-proteomic technology for analysis of proteins in small volumes of respiratory mucosal samples. Furthermore, when we compared nasal wash samples from influenza-infected patients with viral load ≥ 28 and increased IL-6 and CXCL10 to healthy controls, we identified 162 differentially-expressed proteins between the two groups. This number greatly exceeds the number of DEPs identified in previous studies in human influenza patients. Most of the identified proteins were associated with the host immune response to infection, and changes in protein levels of 151 of the DEPs were significantly correlated with viral load. Most important, SOMAscan identified differentially expressed proteins heretofore not associated with respiratory influenza infection in humans. Our study is the first report for the use of SOMAscan to screen nasal secretions. It establishes a precedent for micro-proteomic quantification of proteins that reflect ongoing response to respiratory infection.
    • RNAseq expression analysis of resistant and susceptible mice after influenza A virus infection identifies novel genes associated with virus replication and important for host resistance to infection.

      Wilk, Esther; Pandey, Ashutosh K; Leist, Sarah Rebecca; Hatesuer, Bastian; Preusse, Matthias; Pommerenke, Claudia; Wang, Junxi; Schughart, Klaus; Helmholtz Centre for Infection research, Inhoffenstr. 7, D-38124 Braunschweig, Germany. (2015)
      The host response to influenza A infections is strongly influenced by host genetic factors. Animal models of genetically diverse mouse strains are well suited to identify host genes involved in severe pathology, viral replication and immune responses. Here, we have utilized a dual RNAseq approach that allowed us to investigate both viral and host gene expression in the same individual mouse after H1N1 infection.
    • Schistosoma co-infection protects against brain pathology but does not prevent severe disease and death in a murine model of cerebral malaria.

      Bucher, Kirsten; Dietz, Klaus; Lackner, Peter; Pasche, Bastian; Fendel, Rolf; Mordmüller, Benjamin; Ben-Smith, Anne; Hoffmann, Wolfgang H; Institute of Tropical Medicine, University of Tübingen, Tübingen, Germany. (2011-01)
      Co-infections of helminths and malaria parasites are common in human populations in most endemic areas. It has been suggested that concomitant helminth infections inhibit the control of malaria parasitemia but down-modulate severe malarial disease. We tested this hypothesis using a murine co-infection model of schistosomiasis and cerebral malaria. C57BL/6 mice were infected with Schistosoma mansoni and 8-9 weeks later, when Schistosoma infection was patent, mice were co-infected with Plasmodium berghei ANKA strain. We found that a concomitant Schistosoma infection increased parasitemia at the beginning of the P. berghei infection. It did not protect against P. berghei-induced weight loss and hypothermia, and P. berghei-mono-infected as well as S. mansoni-P. berghei-co-infected animals showed a high case fatality between days 6 and 8 of malarial infection. However, co-infection significantly reduced P. berghei-induced brain pathology. Over 40% of the S. mansoni-P. berghei-co-infected animals that died during this period were completely protected against haemorrhaging, plugging of blood vessels and infiltration, indicating that mortality in these animals was not related to cerebral disease. Schistosoma mansoni-P. berghei-co-infected mice had elevated plasma concentrations of IL-5 and IL-13 and on day 6 lower levels of IFN-γ, IL-10, monocyte chemoattractant protein-1 (MCP-1) and monokine induced by IFN-γ (MIG) than P. berghei-mono-infected mice. We conclude that in P. berghei infections, disease and early death are caused by distinct pathogenic mechanisms, which develop in parallel and are differentially influenced by the immune response to S. mansoni. This might explain why, in co-infected mice, death could be induced in the absence of brain pathology.
    • Segregation of a spontaneous Klrd1 (CD94) mutation in DBA/2 mouse substrains.

      Shin, Dai-Lun; Pandey, Ashutosh K; Ziebarth, Jesse Dylan; Mulligan, Megan K; Williams, Robert W; Geffers, Robert; Hatesuer, Bastian; Schughart, Klaus; Wilk, Esther; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2015-02)
      Current model DBA/2J (D2J) mice lack CD94 expression due to a deletion spanning the last coding exon of the Klrd1 gene that occurred in the mid- to late 1980s. In contrast, DBA/2JRj (D2Rj) mice, crosses derived from DBA/2J before 1984, and C57BL/6J (B6) mice lack the deletion and have normal CD94 expression. For example, BXD lines (BXD1-32) generated in the 1970s by crossing B6 and D2J do not segregate for the exonic deletion and have high expression, whereas BXD lines 33 and greater were generated after 1990 are segregating for the deletion and have highly variable Klrd1 expression. We performed quantitative trait locus analysis of Klrd1 expression by using BXD lines with different generation times and found that the expression difference in Klrd1 in the later BXD set is driven by a strong cis-acting expression quantitative trait locus. Although the Klrd1/CD94 locus is essential for mousepox resistance, the genetic variation among D2 substrains and the later set of BXD strains is not associated with susceptibility to the Influenza A virus PR8 strain. Substrains with nearly identical genetic backgrounds that are segregating functional variants such as the Klrd1 deletion are useful genetic tools to investigate biological function.
    • Self-collected nasal swabs to detect infection and colonization: a useful tool for population-based epidemiological studies?

      Akmatov, M K; Pessler, F; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2011-09)
      Population-based epidemiological studies on infectious diseases are limited by methodological problems that may not be encountered in other fields of epidemiology. The acute or asymptomatic nature of many infections hinders a timely diagnosis by trained personnel in a study centre, indicating the need for new collection methods of biological specimens. One alternative approach is to have the participants collect the specimens themselves, for instance nasal swabs for the detection of bacterial or viral pathogens. Although self-collection is widely accepted in clinical studies of specific populations (e.g., self-collection of vaginal swabs by young women to diagnose sexually transmitted infections), it has not been employed much in population-based studies. Here, we review recent experience with self-collection of nasal swabs for the detection of microorganisms and discuss future prospects and applications for this technique.
    • Sustained viral load and late death in Rag2-/- mice after influenza A virus infection

      Wu, Haiya; Haist, Verena; Baumgärtner, Wolfgang; Schughart, Klaus; Helmholtz Centre for infection research, Ihoffenstr. 7, 38124 Braunschweig, Germany. (2010-07-28)
      Abstract The importance of the adaptive immune response for secondary influenza infections and protection from a lethal challenge after vaccination has been well documented. However, some controversy still exists concerning the specific involvement of B and T cells during a primary infection. Here, we have followed the survival, weight loss, viral load and lung pathology in Rag2 -/- knock-out mice after infection with influenza A virus (H1N1). Infected wild type mice initially lost weight early after infection but then cleared the virus and recovered. Rag2 -/- mice, however, showed similar weight loss kinetics in the early stages after infection but weight loss continued post infection and culminated in death. In contrast to wild type mice, Rag2 -/- mice were not able to clear the virus, despite an increased inflammatory response. Furthermore, they did not recruit virus-specific lymphocytes into the lung in the later stages after infection and exhibited sustained pulmonary lesions.
    • SYSGENET: a meeting report from a new European network for systems genetics.

      Schughart, Klaus; Department of Infection Genetics, Helmholtz Centre for Infection Research, 38124, Braunschweig, Germany, klaus.schughart@helmholtz-hzi.de. (2010-07-11)
      The first scientific meeting of the newly established European SYSGENET network took place at the Helmholtz Centre for Infection Research (HZI) in Braunschweig, April 7-9, 2010. About 50 researchers working in the field of systems genetics using mouse genetic reference populations (GRP) participated in the meeting and exchanged their results, phenotyping approaches, and data analysis tools for studying systems genetics. In addition, the future of GRP resources and phenotyping in Europe was discussed.
    • Systems genetics of liver fibrosis: identification of fibrogenic and expression quantitative trait loci in the BXD murine reference population.

      Hall, Rabea A; Liebe, Roman; Hochrath, Katrin; Kazakov, Andrey; Alberts, Rudi; Laufs, Ulrich; Böhm, Michael; Fischer, Hans-Peter; Williams, Robert W; Schughart, Klaus; Weber, Susanne N; Lammert, Frank (2014)
      The progression of liver fibrosis in response to chronic injury varies considerably among individual patients. The underlying genetics is highly complex due to large numbers of potential genes, environmental factors and cell types involved. Here, we provide the first toxicogenomic analysis of liver fibrosis induced by carbon tetrachloride in the murine 'genetic reference panel' of recombinant inbred BXD lines. Our aim was to define the core of risk genes and gene interaction networks that control fibrosis progression. Liver fibrosis phenotypes and gene expression profiles were determined in 35 BXD lines. Quantitative trait locus (QTL) analysis identified seven genomic loci influencing fibrosis phenotypes (pQTLs) with genome-wide significance on chromosomes 4, 5, 7, 12, and 17. Stepwise refinement was based on expression QTL mapping with stringent selection criteria, reducing the number of 1,351 candidate genes located in the pQTLs to a final list of 11 cis-regulated genes. Our findings demonstrate that the BXD reference population represents a powerful experimental resource for shortlisting the genes within a regulatory network that determine the liver's vulnerability to chronic injury.
    • TLR8 regulation of LILRA3 in monocytes is abrogated in human immunodeficiency virus infection and correlates to CD4 counts and virus loads.

      Low, Hui Zhi; Ahrenstorf, Gerrit; Pommerenke, Claudia; Habermann, Nadine; Schughart, Klaus; Ordóñez, David; Stripecke, Renata; Wilk, Esther; Witte, Torsten; Department of Clinical Immunology and Rheumatology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625, Hannover, Germany. (2016)
      LILRA3 is an immunostimulatory molecule which can conditionally induce the proliferation of cytotoxic cells. LILRA3 has a deletion genotype which is associated with multiple immune disorders. In this study, we wanted to analyze the regulation of LILRA3 and its significance in the context of HIV infection.
    • TMPRSS11A activates the influenza A virus hemagglutinin and the MERS coronavirus spike protein and is insensitive against blockade by HAI-1.

      Zmora, Pawel; Hoffmann, Markus; Kollmus, Heike; Moldenhauer, Anna-Sophie; Danov, Olga; Braun, Armin; Winkler, Michael; Schughart, Klaus; Pöhlmann, Stefan; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-09-07)
      The influenza virus hemagglutinin (HA) facilitates viral entry into target cells. Cleavage of HA by host cell proteases is essential for viral infectivity, and the responsible enzymes are potential targets for antiviral intervention. The type II transmembrane serine protease (TTSP) TMPRSS2 has been identified as an HA activator in cell culture and in the infected host. However, it is less clear whether TMPRSS2-related enzymes can also activate HA for spread in target cells. Moreover, the activity of cellular serine protease inhibitors against HA-activating TTSPs is poorly understood. Here, we show that TMPRSS11A, another member of the TTSP family, cleaves and activates the influenza A virus (FLUAV) HA and the Middle East respiratory syndrome coronavirus spike protein (MERS-S). Moreover, we demonstrate that TMPRSS11A is expressed in murine tracheal epithelium, which is a target of FLUAV infection, and in human trachea, suggesting that the protease could support FLUAV spread in patients. Finally, we show that HA activation by the TMPRSS11A-related enzymes human airway tryptase and DESC1, but not TMPRSS11A itself, is blocked by the cellular serine protease inhibitor hepatocyte growth factor activator inhibitor type-1 (HAI-1). Our results suggest that TMPRSS11A could promote FLUAV spread in target cells and that HA-activating TTSPs exhibit differential sensitivity to blockade by cellular serine protease inhibitors.
    • Tmprss2 Is Essential for Influenza H1N1 Virus Pathogenesis in Mice

      Hatesuer, Bastian; Bertram, Stephanie; Mehnert, Nora; Bahgat, Mahmoud M.; Nelson, Peter S.; Pöhlman, Stefan; Schughart, Klaus; Basler, Christopher F.; Infectiongenetics, Helmholtz Centre for Infection research, Inhoffenstr. 7, D38124 Braunschweig, Germany (2013-12-10)