Leibniz Center for Medicine and Biosciences

Borstel-Hohenraden, Germany

Leibniz Center for Medicine and Biosciences

Borstel-Hohenraden, Germany
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Yildirim A.O.,University of Marburg | Yildirim A.O.,Institute of Lung Biology and Disease | Muyal V.,University of Marburg | John G.,Justus Liebig University | And 5 more authors.
American Journal of Respiratory and Critical Care Medicine | Year: 2010

Rationale: Emphysema is characterized by destruction of alveoli with ensuing airspace enlargement and loss of alveoli. Induction of alveolar regeneration is still a major challenge in emphysema therapy. Objectives: To investigate whether therapeutic application of palifermin (ΔN23-KGF) is able to induce a regenerative response in distal lungparenchyma after induction of pulmonaryemphysema. Methods: Mice were therapeutically treated at three occasions by oropharyngeal aspiration of 10 mg ΔN23-KGF per kg body weight after induction of emphysema by porcine pancreatic elastase. Measurements and Main Results: Airflow limitation associated with emphysema was largely reversed as assessed by noninvasive head-out body plethysmography. Porcine pancreatic elastase-induced airspace enlargement and loss of alveoli were partially reversed as assessed by design-based stereology. ΔN23-KGF induced proliferation of epithelium, endothelium, and fibroblasts being associated with enhanceddifferentiation as well as increased expression of vascular endothelial growth factor, vascular endothelial growth factor receptors, transforming growth factor (TGF)-β1, TGF-β2, (phospho-) Smad2, plasminogen activator inhibitor-1, and elastin as assessed by quantitative reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry. ΔN23-KGF induced the expression of TGF-β1 in and release of active TGF-β1 from primary mouse alveolar epithelial type 2 (AE2) cells, murine AE2-like cells LA-4, and cocultures of LA-4 and murine lung fibroblasts (MLF), but not in MLF cultured alone. Recombinant TGF-β1 but not ΔN23-KGF induced elastin gene expression in MLF. Blockade of TGF-signaling by neutralizing antibody abolished these effects of ΔN23-KGF in LA-4/MLF cocultures. Conclusions: Our data demonstrate that therapeutic application of ΔN23-KGF has the potential to induce alveolar maintenance programs in emphysematous lungs and suggest that the regenerative effect on interstitial tissue is linked to AE2 cell-derived TGF-β1.

Suomalainen M.,University of Helsinki | Lobo L.A.,University of Helsinki | Brandenburg K.,Leibniz Center for Medicine and Biosciences | Lindner B.,Leibniz Center for Medicine and Biosciences | And 5 more authors.
Infection and Immunity | Year: 2010

The Pla surface protease of Yersinia pestis activates human plasminogen and is a central virulence factor in bubonic and pneumonic plague. Pla is a transmembrane β-barrel protein and member of the omptin family of outer membrane proteases which require bound lipopolysaccharide (LPS) to be proteolytically active. Plasminogen activation and autoprocessing of Pla were dramatically higher in Y. pestis cells grown at 37°C than in cells grown at 20°C; the difference in enzymatic activity by far exceeded the increase in the cellular content of the Pla protein. Y. pestis modifies its LPS structure in response to growth temperature. We purified His6-Pla under denaturing conditions and compared various LPS types for their capacity to enhance plasmin formation by His6-Pla solubilized in detergent. Reactivation of His6-Pla was higher with Y. pestis LPSs isolated from bacteria grown at 37°C than with LPSs from cells grown at 25°C. Lack of O antigens and the presence of the outer core region as well as a lowered level of acylation in LPS were found to enhance the Pla-LPS interaction. Genetic substitution of arginine 138, which is part of a three-dimensional protein motif for binding to lipid A phosphates, decreased both the enzymatic activity of His6-Pla and the amount of Pla in Y. pestis cells, suggesting the importance of the Pla-lipid A phosphate interaction. The temperature-induced changes in LPS are known to help Y. pestis to avoid innate immune responses, and our results strongly suggest that they also potentiate Pla-mediated proteolysis. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Polansky J.K.,Helmholtz Center for Infection Research | Polansky J.K.,Leibniz Center for Medicine and Biosciences | Schreiber L.,Helmholtz Center for Infection Research | Thelemann C.,Charité - Medical University of Berlin | And 7 more authors.
Journal of Molecular Medicine | Year: 2010

The forkhead-box protein P3 (Foxp3) is a key transcription factor for the development and suppressive activity of regulatory T cells (Tregs), a T cell subset critically involved in the maintenance of self-tolerance and prevention of over-shooting immune responses. However, the transcriptional regulation of Foxp3 expression remains incompletely understood. We have previously shown that epigenetic modifications in the CpG-rich Treg-specific demethylated region (TSDR) in the Foxp3 locus are associated with stable Foxp3 expression. We now demonstrate that the methylation state of the CpG motifs within the TSDR controls its transcriptional activity rather than a Treg-specific transcription factor network. By systematically mutating every CpG motif within the TSDR, we could identify four CpG motifs, which are critically determining the transcriptional activity of the TSDR and which serve as binding sites for essential transcription factors, such as CREB/ATF and NF-κB, which have previously been shown to bind to this element. The transcription factor Ets-1 was here identified as an additional molecular player that specifically binds to the TSDR in a demethylation-dependent manner in vitro. Disruption of the Ets-1 binding sites within the TSDR drastically reduced its transcriptional enhancer activity. In addition, we found Ets-1 bound to the demethylated TSDR in ex vivo isolated Tregs, but not to the methylated TSDR in conventional CD4+ T cells. We therefore propose that Ets-1 is part of a larger protein complex, which binds to the TSDR only in its demethylated state, thereby restricting stable Foxp3 expression to the Treg lineage. © 2010 The Author(s).

Mertens K.,Leibniz Center for Medicine and Biosciences | Muller-Loennies S.,Leibniz Center for Medicine and Biosciences | Stengel P.,University of Lübeck | Podschun R.,University of Kiel | And 2 more authors.
Innate Immunity | Year: 2010

Raoultella terrigena ATCC 33257, recently reclassified from the genus Klebsiella, is a drinking water isolate and belongs to a large group of non-typeable Klebsiella and Raoultella strains. Using an O-antiserum against a capsule-deficient mutant of this strain, we could show a high prevalence (10.5%) of the R. terrigena O-serotype among non-typeable, clinical Klebsiella and Raoultella isolates. We observed a strong serological cross-reaction with the K. pneumoniae O12 reference strain, indicating that a large percentage of these non-typeable strains may belong to the O12 serotype, although these are currently not detectable by the K. pneumoniae O12 reference antiserum in use. Therefore, we analyzed the O-polysaccharide (O-PS) structure and genetic organization of the wb gene cluster of R. terrigena ATCC 33257, and both confirmed a close relation of R. terrigena and K. pneumoniae O12. The two strains possess an identical O-PS, lipopolysaccharide core structure, and genetic organization of the wb gene cluster. Heterologous expression of the R. terrigena wb gene cluster in Escherichia coli K-12 resulted in the WecA-dependent synthesis of an O-PS reactive with the K. pneumoniae O12 antiserum. The serological data presented here suggest a higher prevalence of the O12-serotype among Klebsiella and Raoultella isolates than generally assumed. © SAGE Publications 2010.

Fischer N.,University of Hamburg | Schulz C.,University of Hamburg | Stieler K.,University of Hamburg | Hohn O.,Robert Koch Institute | And 3 more authors.
Emerging Infectious Diseases | Year: 2010

Xenotropic murine leukemia virus-related gammaretrovirus (XMRV) has been recently associated with prostate cancer and chronic fatigue syndrome. To identify nucleic acid sequences, we examined respiratory secretions by using PCR. XMRV-specifi c sequences were detected in 2%-3% of samples from 168 immunocompetent carriers and ≈10% of samples from 161 immunocompromised patients.

Posch G.,University of Vienna | Andrukhov O.,Medical University of Vienna | Vinogradov E.,NRC Institute for Biological Sciences | Lindner B.,Leibniz Center for Medicine and Biosciences | And 3 more authors.
Clinical and Vaccine Immunology | Year: 2013

Tannerella forsythia is a Gram-negative anaerobic organism that inhabits subgingival plaque biofilms and is covered with a so far unique surface layer composed of two glycoproteins. It belongs to the so-called "red complex" of bacteria comprising species that are associated with periodontal disease. While the surface layer glycoprotein glycan structure had been elucidated recently and found to be a virulence factor, no structural data on the lipopolysaccharide (LPS) of this organism were available. In this study, the T. forsythia LPS structure was partially elucidated by a combined mass spectrometry (MS) and nuclear magnetic resonance spectroscopy (NMR) approach and initial experiments to characterize its immunostimulatory potential were performed. The T. forsythia LPS is a complex, rough-type LPS with a core region composed of one 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) residue, three mannose residues, and two glucosamine residues. MS analyses of O-deacylated LPS proved that, in addition, one phosphoethanolamine residue and most likely one galactose-phosphate residue were present, however, their positions could not be identified. Stimulation of human macrophages with T. forsythia LPS resulted in the production of the proinflammatory cytokines interleukin-1 (IL-1), IL-6, and tumor necrosis factor alpha in a dose-dependent manner. The response to T. forsythia LPS was observed only upon stimulation in the presence of fetal calf serum (FCS), whereas no cytokine production was observed in the absence of FCS. This finding suggests that the presence of certain additional cofactors is crucial for the immune response induced by T. forsythia LPS. Copyright © 2013, American Society for Microbiology. All Rights Reserved.

Claes A.-K.,University of Toronto | Claes A.-K.,Leibniz Center for Medicine and Biosciences | Claes A.-K.,University of Kiel | Zhou J.Y.,University of Toronto | Philpott D.J.,University of Toronto
Physiology | Year: 2015

The NOD-like receptors (NLRs) are cytosolic pattern-recognition receptors, which are critically involved in mucosal immune defense. The association of the NLR, NOD2, with inflammatory bowel disease first pointed to the NLRs potential function as guardians of the intestinal barrier. Since then, several studies have emphasized the importance of NLRs in maintaining gut homeostasis and intestinal infections, and in shaping the microbiota. In this review, we will highlight the function of NLRs in intestinal inflammation. © 2015 Int.

Schromm A.B.,Emmy Noether Group of Immunobiophysics | Reiling N.,Leibniz Center for Medicine and Biosciences | Howe J.,Leibniz Center for Medicine and Biosciences | Wiesmuller K.-H.,EMC Microcollections GmbH | And 2 more authors.
Innate Immunity | Year: 2010

The innate immune response provides a critical first-line defense against Mycobacterium tuberculosis, an intracellular pathogen that represents a major health threat world-wide. A synthetic lipopeptide (LP) mimicking the lipid moiety of the cell-wall associated 19-kDa lipoprotein from M. tuberculosis has recently been assigned an important role in the induction of an antibacterial immune response in host macrophages. Here, we present experimental data on the biological activities and the biophysical mechanisms underlying cell activation by synthetic 19-kDa M. tuberculosis-derived lipopeptide (Mtb-LP). Investigation of the geometry of the LP (i.e. the molecular conformation and supramolecular aggregate structure) and the preference for membrane intercalation provide an explanation for the biological activities of the mycobacterial LP. Cell activation by low concentrations of Mtb-LP was enhanced by the lipopolysaccharideĝ€"binding protein and CD14. However, surprisingly, we found that activation of human macrophages to induce pro- as well as antiinflammatory mediators (tumor necrosis factor(TNF)-±, Interleukin(IL)-6, IL-8, and IL-10) in response to the Mtb-LP is strongly reduced in the presence of serum. This observation could be confirmed for the immune response of murine macrophages which showed a strongly enhanced TNF-± release in the absence of serum, suggesting that the molecular mechanisms of immune recognition of the Mtb-LP are tailored to the ambient conditions of the lung. © 2010 SAGE Publications.

PubMed | Leiden University and Leibniz Center for Medicine and Biosciences
Type: Journal Article | Journal: Glycobiology | Year: 2015

During the complex lifecycle of Schistosoma mansoni, a large variety of glycans is expressed. To many of these glycans, antibodies are induced by the infected host and some might be targets for vaccines or diagnostic tests. Spatial changes in glycan expression during schistosome development are largely unexplored. To study the surface-exposed glycans during the important initial stages of infection, we analyzed the binding of a panel of anti-glycan monoclonal antibodies (mAbs) to cercariae and schistosomula up to 72 h after transformation by immunofluorescence microscopy. The mAb specificity toward their natural targets was studied using a microarray containing a wide range of schistosomal N-glycans, O-glycans and glycosphingolipid glycans. With the exception of GalNAc1-4(Fuc1-3)GlcNAc (LDN-F), mono- and multifucosylated GalNAc1-4GlcNAc (LDN)-motifs were exposed at the surface of all developmental stages studied. Multifucosylated LDN-motifs were present on cercarial glycocalyx-derived O-glycans as well as cercarial glycolipids. In contrast, the Gal1-4(Fuc1-3)GlcNAc (Lewis X) and LDN-F-motifs, also expressed on cercarial glycolipids, and in addition on a range of cercarial N- and O-glycans, became surface expressed only after transformation of cercariae to schistosomula. In line with the documented shedding of the O-glycan-rich cercarial glycocalyx after transformation these observations suggest that surface accessible multifucosylated LDN-motifs are mostly expressed by O-glycans in cercariae, but principally by glycosphingolipids in schistosomula. We hypothesize that these temporal changes in surface exposure of glycan antigens are relevant to the interaction with the host during the initial stages of infection with schistosomes and discuss the potential of these glycan antigens as intervention targets.

PubMed | Aix - Marseille University, University of Navarra, University of Stockholm and Leibniz Center for Medicine and Biosciences
Type: Journal Article | Journal: The Journal of biological chemistry | Year: 2016

The structures of the lipooligosaccharides fromBrucella melitensismutants affected in the WbkD and ManBcoreproteins have been fully characterized using NMR spectroscopy. The results revealed that disruption ofwbkDgives rise to a rough lipopolysaccharide (R-LPS) with a complete core structure (-d-Glcp-(14)--Kdop-(24)[-d-GlcpN-(16)--d-GlcpN-(14)[-d-GlcpN-(16)]--d-GlcpN-(13)--d-Manp-(15)]--Kdop-(26)--d-GlcpN3N4P-(16)--d-GlcpN3N1P), in addition to components lacking one of the terminal -d-GlcpN and/or the -d-Glcpresidues (48 and 17%, respectively). These structures were identical to those of the R-LPS fromB. melitensisEP, a strain simultaneously expressing both smooth and R-LPS, also studied herein. In contrast, disruption ofmanBcoregives rise to a deep-rough pentasaccharide core (-d-Glcp-(14)--Kdop-(24)--Kdop-(26)--d-GlcpN3N4P-(16)--d-GlcpN3N1P) as the major component (63%), as well as a minor tetrasaccharide component lacking the terminal -d-Glcpresidue (37%). These results are in agreement with the predicted functions of the WbkD (glycosyltransferase involved in the biosynthesis of the O-antigen) and ManBcoreproteins (phosphomannomutase involved in the biosynthesis of a mannosyl precursor needed for the biosynthesis of the core and O-antigen). We also report that deletion ofB. melitensis wadCremoves the core oligosaccharide branch not linked to the O-antigen causing an increase in overall negative charge of the remaining LPS inner section. This is in agreement with the mannosyltransferase role predicted for WadC and the lack of GlcpN residues in the defective core oligosaccharide. Despite carrying the O-antigen essential inB. melitensisvirulence, the core deficiency in thewadCmutant structure resulted in a more efficient detection by innate immunity and attenuation, proving the role of the -d-GlcpN-(16)--d-GlcpN-(14)[-d-GlcpN-(16)]--d-GlcpN-(13)--d-Manp-(15) structure in virulence.

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