Dokument-ID Dokumenttyp Verfasser/Autoren Herausgeber Haupttitel Abstract Auflage Verlagsort Verlag Erscheinungsjahr Seitenzahl Schriftenreihe Titel Schriftenreihe Bandzahl ISBN Quelle der Hochschulschrift Konferenzname Quelle:Titel Quelle:Jahrgang Quelle:Heftnummer Quelle:Erste Seite Quelle:Letzte Seite URN DOI Abteilungen OPUS4-19668 Wissenschaftlicher Artikel Ono, Mitsuaki; Sonoyama, Wataru; Nema, Kazuki; Hara, Emilio Satoshi; Oida, Yasutaka; Pham, Hai Thanh; Yamamoto, Katushi; Hirota, Kazuo; Sugama, Kazushige; Sebald, Walter; Kuboki, Takuo Regeneration of calvarial defects with Escherichia coli-derived rhBMP-2 adsorbed in PLGA membrane Objective: Escherichia coli-derived recombinant human bone morphogenetic protein-2 (E-BMP-2) has been shown to be as effective as mammalian cell-derived BMP-2. However, several in vitro and in vivo experiments are still necessary to validate the effectiveness of E-BMP-2 due to the difference in synthesis process, mainly related to protein nonglycosylation. The objective of this study was to investigate whether biodegradable polylactide-co-glycolide (PLGA) membrane is a suitable carrier for E-BMP-2 delivery for bone regeneration of critical-sized defects in rat calvaria. Materials and Methods: First, the osteoinductive effect of E-BMP-2 was confirmed in vitro in mouse bone marrow stromal cells by analysis of osteocalcin mRNA levels, and calcium deposition was detected by alizarin red staining. Before in vivo experiments, the release profile of E-BMP-2 from PLGA membranes was determined by ELISA. E-BMP-2 (0, 1, 5 and 10 μg/μl) was applied for ectopic and orthotopic bone formation and was analyzed by X-ray, micro-CT and histology. Results: Release-profile testing showed that PLGA membrane could retain 94% of the initially applied E-BMP-2. Ectopic bone formation assay revealed that combination of E-BMP-2/PLGA membrane strongly induced bone formation. Stronger osteoinductivity with complete repair of critical-sized defects was observed only with PLGA membranes adsorbed with 5 and 10 μg/μl of E-BMP-2, whereas no bone formation was observed in the groups that received no membrane or 0-μg/μl dose of E-BMP-2. Conclusion: PLGA membrane was shown to be a suitable carrier for sustained release of E-BMP-2, and the E-BMP-2/PLGA membrane combination was demonstrated to be efficient in bone regeneration in a model of critical-sized defects. 2014 9 Cells Tissues Organs 198 5 367 376 urn:nbn:de:bvb:20-opus-196680 10.1159/000356947 Theodor-Boveri-Institut für Biowissenschaften OPUS4-14018 Wissenschaftlicher Artikel Seher, Axel; Nickel, Joachim; Mueller, Thomas D.; Kneitz, Susanne; Gebhardt, Susanne; Meyer ter Vehn, Tobias; Schlunck, Guenther; Sebald, Walter Gene expression profiling of connective tissue growth factor (CTGF) stimulated primary human tenon fibroblasts reveals an inflammatory and wound healing response in vitro Purpose: The biologic relevance of human connective tissue growth factor (hCTGF) for primary human tenon fibroblasts (HTFs) was investigated by RNA expression profiling using affymetrix (TM) oligonucleotide array technology to identify genes that are regulated by hCTGF. Methods: Recombinant hCTGF was expressed in HEK293T cells and purified by affinity and gel chromatography. Specificity and biologic activity of hCTGF was confirmed by biosensor interaction analysis and proliferation assays. For RNA expression profiling HTFs were stimulated with hCTGF for 48h and analyzed using affymetrix (TM) oligonucleotide array technology. Results were validated by real time RT-PCR. Results: hCTGF induces various groups of genes responsible for a wound healing and inflammatory response in HTFs. A new subset of CTGF inducible inflammatory genes was discovered (e.g., chemokine [C-X-C motif] ligand 1 [CXCL1], chemokine [C-X-C motif] ligand 6 [CXCL6], interleukin 6 [IL6], and interleukin 8 [IL8]). We also identified genes that can transmit the known biologic functions initiated by CTGF such as proliferation and extracellular matrix remodelling. Of special interest is a group of genes, e.g., osteoglycin (OGN) and osteomodulin (OMD), which are known to play a key role in osteoblast biology. Conclusions: This study specifies the important role of hCTGF for primary tenon fibroblast function. The RNA expression profile yields new insights into the relevance of hCTGF in influencing biologic processes like wound healing, inflammation, proliferation, and extracellular matrix remodelling in vitro via transcriptional regulation of specific genes. The results suggest that CTGF potentially acts as a modulating factor in inflammatory and wound healing response in fibroblasts of the human eye. 2011 53-62 Molecular Vision 17 08. Okt urn:nbn:de:bvb:20-opus-140189 Augenklinik und Poliklinik OPUS4-15814 Wissenschaftlicher Artikel Seher, Axel; Lagler, Charlotte; Stühmer, Thorsten; Müller-Richter, Urs Dietmar Achim; Kübler, Alexander Christian; Sebald, Walter; Müller, Thomas Dieter; Nickel, Joachim Utilizing BMP-2 muteins for treatment of multiple myeloma Multiple myeloma (MM) represents a haematological cancer characterized by the pathological hyper proliferation of antibody-producing B-lymphocytes. Patients typically suffer from kidney malfunction and skeletal disorders. In the context of MM, the transforming growth factor β (TGFβ) member Activin A was recently identified as a promoter of both accompanying symptoms. Because studies have shown that bone morphogenetic protein (BMP)-2-mediated activities are counteracted by Activin A, we analysed whether BMP2, which also binds to the Activin A receptors ActRII and ActRIIB but activates the alternative SMAD-1/5/8 pathway, can be used to antagonize Activin A activities, such as in the context of MM. Therefore three BMP2 derivatives were generated with modified binding activities for the type II (ActRIIB) and/or type I receptor (BMPRIA) showing either increased or decreased BMP2 activity. In the context of MM these BMP2 muteins show two functionalities since they act as a) an anti-proliferative/apoptotic agent against neoplastic B-cells, b) as a bone-formation promoting growth factor. The molecular basis of both activities was shown in two different cellular models to clearly rely on the properties of the investigated BMP2 muteins to compete for the binding of Activin A to the Activin type II receptors. The experimental outcome suggests new therapeutic strategies using BMP2 variants in the treatment of MM-related pathologies. 2017 e0174884 PLoS ONE 12 5 urn:nbn:de:bvb:20-opus-158144 10.1371/journal.pone.0174884 Theodor-Boveri-Institut für Biowissenschaften OPUS4-12555 Wissenschaftlicher Artikel Klammert, Uwe; Müller, Thomas D.; Hellmann, Tina V.; Wuerzler, Kristian K.; Kotzsch, Alexander; Schliermann, Anna; Schmitz, Werner; Kuebler, Alexander C.; Sebald, Walter; Nickel, Joachim GDF-5 can act as a context-dependent BMP-2 antagonist Background Bone morphogenetic protein (BMP)-2 and growth and differentiation factor (GDF)-5 are two related transforming growth factor (TGF)-β family members with important functions in embryonic development and tissue homeostasis. BMP-2 is best known for its osteoinductive properties whereas GDF-5—as evident from its alternative name, cartilage derived morphogenetic protein 1—plays an important role in the formation of cartilage. In spite of these differences both factors signal by binding to the same subset of BMP receptors, raising the question how these different functionalities are generated. The largest difference in receptor binding is observed in the interaction with the type I receptor BMPR-IA. GDF-5, in contrast to BMP-2, shows preferential binding to the isoform BMPR-IB, which is abrogated by a single amino acid (A57R) substitution. The resulting variant, GDF-5 R57A, represents a "BMP-2 mimic" with respect to BMP receptor binding. In this study we thus wanted to analyze whether the two growth factors can induce distinct signals via an identically composed receptor. Results Unexpectedly and dependent on the cellular context, GDF-5 R57A showed clear differences in its activity compared to BMP-2. In ATDC-5 cells, both ligands induced alkaline phosphatase (ALP) expression with similar potency. But in C2C12 cells, the BMP-2 mimic GDF-5 R57A (and also wild-type GDF-5) clearly antagonized BMP-2-mediated ALP expression, despite signaling in both cell lines occurring solely via BMPR-IA. The BMP-2- antagonizing properties of GDF-5 and GDF-5 R57A could also be observed in vivo when implanting BMP-2 and either one of the two GDF-5 ligands simultaneously at heterotopic sites. Conclusions Although comparison of the crystal structures of the GDF-5 R57A:BMPR-IAEC- and BMP-2:BMPR-IAEC complex revealed small ligand-specific differences, these cannot account for the different signaling characteristics because the complexes seem identical in both differently reacting cell lines. We thus predict an additional component, most likely a not yet identified GDF-5-specific co-receptor, which alters the output of the signaling complexes. Hence the presence or absence of this component then switches GDF-5′s signaling capabilities to act either similar to BMP-2 or as a BMP-2 antagonist. These findings might shed new light on the role of GDF-5, e.g., in cartilage maintenance and/or limb development in that it might act as an inhibitor of signaling events initiated by other BMPs. 2015 BMC Biology 13 77 urn:nbn:de:bvb:20-opus-125550 10.1186/s12915-015-0183-8 Klinik und Poliklinik für Mund-, Kiefer- und Plastische Gesichtschirurgie OPUS4-7222 Wissenschaftlicher Artikel Sebald, Walter; Arends, Hermann; McCarthy, John E. G. Isolation and manipulation of genes coding for energy-transducing enzymes from Neurospora crassa and Escherichia coli No abstract available. 1984 urn:nbn:de:bvb:20-opus-86768 Theodor-Boveri-Institut für Biowissenschaften OPUS4-7221 Wissenschaftlicher Artikel Duschl, Albert; Jahn, Ute; Bertling, Claudia; Sebald, Walter A comparison of assays for the response of primary human T-cells upon stimulation with interleukin-2, interleukin-4 and interleukin-7 The most commonly used assay to quantitate the response of peripheral T~cells upon stimulation with growth factors is determination of incorporated (JH]TdR. We compared thls test to three other methods: 1. direct countlog of cells with a Coulter type counter as reference assay, 2. a colorimetric assay using the tetrazolium dye 3-[ 4,S-dimethylthiazol-l-yl]-2,5diphenyl tetrazolium (MTT), which is a cheap and increasingly popular non-radioactive method and 3. incorporation of the thymidine analog 5-bromo-2'-deoxyuridine detection with a monoclonal antibody on cytospins. Primary human PHA-blasts from >30 healthy individuals were stimulated with IL-2, IL-4 aod IL-7 and assayed with up to four different methods. We discuss the advantages and disadvantages of the assays used and tbe effects of differences between cell preparations. We observed no significant variations between individuals for the dose dependence, but the relative emctency of IL4 compared to IL-2 and IL-7 was variable. This was probably due to the slower response observed upon stimulation with this factor. 1992 urn:nbn:de:bvb:20-opus-86750 Theodor-Boveri-Institut für Biowissenschaften OPUS4-7167 Wissenschaftlicher Artikel Kübler, Norbert; Reuther, Jürgen; Kirchner, Thomas; Priessnitz, Bernd; Sebald, Walter Osteoinductive, morphologic, and biomechanical properties of autolyzed, antigen-extracted, allogeneic human bone Autolyzed, antigen-extracted, allogeneic (AAA) bone was prepared from human cortical bone and its morphologic, biomechanical, and osteoinductive properlies were compared with untreated (frozen) as well as lyophilized human bone. Scanning electron microscopy revealed removal of inprganic calcium phosphates and persistence of shrunken collagen fibrils on the surface of AAA bone matrix. Biomechanical testing of differently prepared bone samples showed that lyophilization increased both the modulus of elasticity (P < .00001) and the compressive strength (P < .00001 ). Depending on the depth of decalcification in the preparation of AAA bone, both measured values decreased in rehydrated AAA bone compared with untreated bone {P < .00001 ). Completely demineralized and rehydrated AAA bone was soft, flexible, and showed very little compressive strength. Differences in biomechanical behavior between samples drilled longitudinally or perpendicularly to the diaphyseal bone axis were observed. Xenogeneic human bone samples were implanted in muscle pouches of Sprague-Dawley rats for 6 weeks. AAA bone implants showed chondrogenesis and osteogenesis in 50% of the cases, while untreated or lyophilized bone implants induced no new cartilage or bone formation. As decalcification exposed xenogeneic organic matrix components, AAA bone implants provoked the highest inflammatory reaction. When AAA bone samples were implanted in immunosuppressed rats, the inflammatory reaction was suppressed and 94o/o of the implants showed endochondral bone formation. The chondroinductivity of the bone samples also was tested in vitro using neonatal rat muscle tissue to avoid interference with inflammatory cells and secreted cytokines. In this assay, 68°/o of AAA bone samples induced chondroneogenesis, while untreated as weil as lyophilized bone samples failed to induce any cartilage formation. The results clearly dernonstrafe that AAA bone has osteoinductive properties. Biomechanical stability of AAA bone implants depends on the degree of demineralization. Thus, they can be prepared in an appropriate manner for different indications in oral and maxillofacial surgery. 1993 urn:nbn:de:bvb:20-opus-86715 Theodor-Boveri-Institut für Biowissenschaften OPUS4-9719 Wissenschaftlicher Artikel Mueller, Thomas D.; Fiebig, Juliane E.; Weidauer, Stella E.; Qiu, Li-Yan; Bauer, Markus; Schmieder, Peter; Beerbaum, Monika; Zhang, Jin-Li; Oschkinat, Hartmut; Sebald, Walter The Clip-Segment of the von Willebrand Domain 1 of the BMP Modulator Protein Crossveinless 2 Is Preformed Bone Morphogenetic Proteins (BMPs) are secreted protein hormones that act as morphogens and exert essential roles during embryonic development of tissues and organs. Signaling by BMPs occurs via hetero-oligomerization of two types of serine/threonine kinase transmembrane receptors. Due to the small number of available receptors for a large number of BMP ligands ligand-receptor promiscuity presents an evident problem requiring additional regulatory mechanisms for ligand-specific signaling. Such additional regulation is achieved through a plethora of extracellular antagonists, among them members of the Chordin superfamily, that modulate BMP signaling activity by binding. The key-element in Chordin-related antagonists for interacting with BMPs is the von Willebrand type C (VWC) module, which is a small domain of about 50 to 60 residues occurring in many different proteins. Although a structure of the VWC domain of the Chordin-member Crossveinless 2 (CV2) bound to BMP-2 has been determined by X-ray crystallography, the molecular mechanism by which the VWC domain binds BMPs has remained unclear. Here we present the NMR structure of the Danio rerio CV2 VWC1 domain in its unbound state showing that the key features for high affinity binding to BMP-2 is a pre-oriented peptide loop. 2013 Molecules urn:nbn:de:bvb:20-opus-97196 10.3390/molecules181011658 Theodor-Boveri-Institut für Biowissenschaften OPUS4-1281 Wissenschaftlicher Artikel Birkmayer, G. D.; Sebald, Walter; Bücher, Th. Cytochrom-Oxydase und ein an diese assoziiertes, markiertes Protein aus 14C-markierten Mitochondrien von Neurospora crassa no abstract available 1969 urn:nbn:de:bvb:20-opus-82135 Theodor-Boveri-Institut für Biowissenschaften OPUS4-2032 Wissenschaftlicher Artikel Schwab, A. J.; Sebald, Walter; Weiss, H. Schnelle Markierung eines mitochondrial synthetisiertem Polypeptids einer Cytochromoxidasen-Präparation aus Neurospora no abstracts available 1972 urn:nbn:de:bvb:20-opus-84206 Theodor-Boveri-Institut für Biowissenschaften