• Antibodies against C-reactive protein cross-react with 60-kilodalton heat shock proteins.

      Udvarnoki, Katalin; Cervenak, László; Uray, Katalin; Hudecz, Ferenc; Kacskovics, Imre; Spallek, Ralf; Singh, Mahavir; Füst, George; Prohászka, Zoltán; Third Department of Medicine, Semmelweis University, H-1125 Budapest, Kútvölgyi st. 4, Hungary. (2007-04)
      C-reactive protein (CRP) is an acute-phase reactant frequently used in histochemistry as a marker of ongoing inflammation. Furthermore, CRP is a powerful biomarker for the prediction of coronary artery disease risk. Heat-shock protein 60 (Hsp60) and CRP are complement-activating molecules, and the effect of their interactions on the regulation of complement activation was studied. However, during the first experiments, we learned that polyclonal anti-CRP antibodies cross-react with Hsp60. Therefore, the aim of our present study was to analyze the cross-reactivity of anti-CRP antibodies (Ab) with Hsp60 in solid-phase enzyme immune assays, in epitope studies using a series of overlapping synthetic peptides, and in Ouchterlony analyses. We found that three different commercial rabbit polyclonal antibodies and two monoclonal (9C9 and CRP-8) anti-CRP antibodies specifically recognize recombinant human Hsp60 and recombinant Mycobacterium tuberculosis Hsp65, respectively. Hsp60 was found to inhibit the binding of anti-CRP polyclonal Ab to Hsp60. Six epitope regions of Hsp60 were recognized by the anti-CRP antibodies, and one region (amino acids [AA] 218 to 232) was recognized by monoclonal antibodies CRP-8 and 9C9. This epitope region of Hsp60 displays 26.6% amino acid identity to CRP AA region 77 to 90. These data suggest that the B-cell epitopes shared between CRP and Hsp60 give rise to a true mimicry-based cross-reaction and the induction of cross-reactive antibodies. Our study underlines the importance of thorough study design and careful interpretation of results while using polyclonal anti-CRP antibodies for histochemistry, especially at low dilutions. Furthermore, analytical interference with Hsp60 in CRP assays should also be tested.
    • Cellular retinaldehyde-binding protein (CRALBP) is a direct downstream target of transcription factor Pax6.

      Boppana, Sridhar; Scheglov, Alexander; Geffers, Robert; Tarabykin, Victor; Max-Planck-Institute for Experimental Medicine, Hermann-Rein Strasse 3, 37075 Göttingen, Germany. boppansr@umdnj.edu (2012-02)
      Transcription factor Pax6 plays an essential role in the expression of other transcription factors, cell adhesion molecules and is crucial for neurogenesis in the developing forebrain. Analysis of gene expression profiles through microarray experiments in Pax6 mutants allowed us to focus on CRALBP, one of the many genes that were downregulated.
    • Deletion of Irf3 and Irf7 Genes in Mice Results in Altered Interferon Pathway Activation and Granulocyte-Dominated Inflammatory Responses to Influenza A Infection.

      Hatesuer, Bastian; Hoang, Hang Thi Thu; Riese, Peggy; Trittel, Stephanie; Gerhauser, Ingo; Elbahesh, Husni; Geffers, Robert; Wilk, Esther; Schughart, Klaus; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017)
      The interferon (IFN) pathway plays an essential role in the innate immune response following viral infections and subsequent shaping of adaptive immunity. Infections with influenza A viruses (IAV) activate the IFN pathway after the recognition of pathogen-specific molecular patterns by respective pattern recognition receptors. The IFN regulatory factors IRF3 and IRF7 are key players in the regulation of type I and III IFN genes. In this study, we analyzed the role of IRF3 and IRF7 for the host response to IAV infections in Irf3-/-, Irf7-/-, and Irf3-/-Irf7-/- knockout mice. While the absence of IRF3 had only a moderate impact on IFN expression, deletion of IRF7 completely abolished IFNα production after infection. In contrast, lack of both IRF3 and IRF7 resulted in the absence of both IFNα and IFNβ after IAV infection. In addition, IAV infection of double knockout mice resulted in a strong increase of mortality associated with a massive influx of granulocytes in the lung and reduced activation of the adaptive immune response.
    • Enantiomer-specific and paracrine leukemogenicity of mutant IDH metabolite 2-hydroxyglutarate.

      Chaturvedi, A; Araujo Cruz, M M; Jyotsana, N; Sharma, A; Goparaju, R; Schwarzer, A; Görlich, K; Schottmann, R; Struys, E A; Jansen, E E; et al. (2016-08)
      Canonical mutations in IDH1 and IDH2 produce high levels of the R-enantiomer of 2-hydroxyglutarate (R-2HG), which is a competitive inhibitor of α-ketoglutarate (αKG)-dependent enzymes and a putative oncometabolite. Mutant IDH1 collaborates with HoxA9 to induce monocytic leukemia in vivo. We used two mouse models and a patient-derived acute myeloid leukemia xenotransplantation (PDX) model to evaluate the in vivo transforming potential of R-2HG, S-2HG and αKG independent of the mutant IDH1 protein. We show that R-2HG, but not S-2HG or αKG, is an oncometabolite in vivo that does not require the mutant IDH1 protein to induce hyperleukocytosis and to accelerate the onset of murine and human leukemia. Thus, circulating R-2HG acts in a paracrine manner and can drive the expansion of many different leukemic and preleukemic clones that may express wild-type IDH1, and therefore can be a driver of clonal evolution and diversity. In addition, we show that the mutant IDH1 protein is a stronger oncogene than R-2HG alone when comparable intracellular R-2HG levels are achieved. We therefore propose R-2HG-independent oncogenic functions of mutant IDH1 that may need to be targeted in addition to R-2HG production to exploit the full therapeutic potential of IDH1 inhibition.
    • Genome-wide localization and expression profiling establish Sp2 as a sequence-specific transcription factor regulating vitally important genes.

      Terrados, Gloria; Finkernagel, Florian; Stielow, Bastian; Sadic, Dennis; Neubert, Juliane; Herdt, Olga; Krause, Michael; Scharfe, Maren; Jarek, Michael; Suske, Guntram; et al. (2012-09)
      The transcription factor Sp2 is essential for early mouse development and for proliferation of mouse embryonic fibroblasts in culture. Yet its mechanisms of action and its target genes are largely unknown. In this study, we have combined RNA interference, in vitro DNA binding, chromatin immunoprecipitation sequencing and global gene-expression profiling to investigate the role of Sp2 for cellular functions, to define target sites and to identify genes regulated by Sp2. We show that Sp2 is important for cellular proliferation that it binds to GC-boxes and occupies proximal promoters of genes essential for vital cellular processes including gene expression, replication, metabolism and signalling. Moreover, we identified important key target genes and cellular pathways that are directly regulated by Sp2. Most significantly, Sp2 binds and activates numerous sequence-specific transcription factor and co-activator genes, and represses the whole battery of cholesterol synthesis genes. Our results establish Sp2 as a sequence-specific regulator of vitally important genes.
    • Hypoxia Enhances Immunosuppression by Inhibiting CD4+ Effector T Cell Function and Promoting Treg Activity.

      Westendorf, Astrid M; Skibbe, Kathrin; Adamczyk, Alexandra; Buer, Jan; Geffers, Robert; Hansen, Wiebke; Pastille, Eva; Jendrossek, Verena; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017)
      Hypoxia occurs in many pathological conditions, including inflammation and cancer. Within this context, hypoxia was shown to inhibit but also to promote T cell responses. Due to this controversial function, we aimed to explore whether an insufficient anti-tumour response during colitis-associated colon cancer could be ascribed to a hypoxic microenvironment.
    • Interleukin-2 improves amyloid pathology, synaptic failure and memory in Alzheimer's disease mice.

      Alves, Sandro; Churlaud, Guillaume; Audrain, Mickael; Michaelsen-Preusse, Kristin; Fol, Romain; Souchet, Benoit; Braudeau, Jérôme; Korte, Martin; Klatzmann, David; Cartier, Nathalie; et al. (2017-03-01)
      Interleukin-2 (IL-2)-deficient mice have cytoarchitectural hippocampal modifications and impaired learning and memory ability reminiscent of Alzheimer's disease. IL-2 stimulates regulatory T cells whose role is to control inflammation. As neuroinflammation contributes to neurodegeneration, we investigated IL-2 in Alzheimer's disease. Therefore, we investigated IL-2 levels in hippocampal biopsies of patients with Alzheimer's disease relative to age-matched control individuals. We then treated APP/PS1ΔE9 mice having established Alzheimer's disease with IL-2 for 5 months using single administration of an AAV-IL-2 vector. We first found decreased IL-2 levels in hippocampal biopsies of patients with Alzheimer's disease. In mice, IL-2-induced systemic and brain regulatory T cells expansion and activation. In the hippocampus, IL-2 induced astrocytic activation and recruitment of astrocytes around amyloid plaques, decreased amyloid-β42/40 ratio and amyloid plaque load, improved synaptic plasticity and significantly rescued spine density. Of note, this tissue remodelling was associated with recovery of memory deficits, as assessed in the Morris water maze task. Altogether, our data strongly suggest that IL-2 can alleviate Alzheimer's disease hallmarks in APP/PS1ΔE9 mice with established pathology. Therefore, this should prompt the investigation of low-dose IL-2 in Alzheimer's disease and other neuroinflammatory/neurodegenerative disorders.