docMolekulare Grundlagen der Regeneration des Skeletts
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Molekulare Grundlagen der Regeneration des Skeletts
Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Molekulare Grundlagen der Regeneration des Skeletts Stefan Mundlos BCRT Institut für Medizinische Genetik, Charité Max-Planck-Institut für Molekulare Genetik Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Wednesday, 30 April 2008 The man who grew a finger By Matthew Price BBC News, Ohio How? Well that's the truly remarkable part. It wasn't a transplant. Mr Spievak re-grew his finger tip. He used a powder - or pixie dust as he sometimes refers to it while telling his story. "There are all sorts of signals in the body," explains Dr Badylak. "I think that within ten years that we will have strategies that will re-grow the bones, and promote the growth of functional tissue around those bones. And that is a major step towards eventually doing the entire limb." That kind of talk has got the US military interested. They are just about to start trials to re-grow parts of the fingers of injured soldiers. 1 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Phasen der Knochenheilung Thrombozyten Hämatom und Entzündung Bildung von neuer Knochenmatrix und Mineralisation Knochen nach „remodeling“ Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies monocytic Progenitors Signalmoleküle Signalmoleküle mesenchymal Progenitors crosstalk KnochenHomöostase lining cells osteocyte Reso rpti on old bone Activation new bone Resorption Formation Mineralization 2 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Komplexe Struktur von trabekulärem Knochen Interaktion von genetischen Programm mit Umwelt - insbesondere mechanische Belastung - Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Biologische Signalmoleküle 3 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Screen for Genes that Control Bone and Cartilage Formation 1) wt 2) 3) 4) ko Compare wt vs mutant mouse Identify regulatory networks Identify genes Identify in callus formation disease genes Bioinformatic analysis of genes and functional testing in vivo chick in vitro mouse Test positive candidates in disease models Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Runx2 Knock out Mouse total absence of bone affected cell types: -osteoblasts -osteoclasts -hypertrophic chondrocytes Wt Runx2 -/- Runx2 essential factor for osteoblast differentiation 4 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Regulation of Transcription Chromatin immunoprecipitation and deep sequencing Identify targets Define regulatory network Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Screen for Genes that Control Bone and Cartilage Formation 1) wt 2) 3) 4) ko Compare wt vs mutant mouse Identify regulatory networks Identify genes Identify in callus formation disease genes Bioinformatic analysis of genes and functional testing in vitro in vivo chick mouse Test positive candidates in disease models 5 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Vorteil: Biomechanik Nachteil: Keine Sequenzen Modellorganismus Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Untersuchung der Frakturheilung am Schaf cDNA-Banken liefern: - Sequenzinformationen - Identifikation aller im Kallus exprimierten Gene 6 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Cluster der qRT-PCR-Ergebnisse Cluster A: 6 von 8 Angiogenese-Genen Cluster B: 13 von 15 ribosomalen Genen Cluster E: 3 der 5 Gene sind Osteoklastenmarker ca. 60% der Kandidaten bisher ohne bekannte spezifische Funktion im Knochen Dr. Peter N. Robinson, Sebastian Bauer Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Screen for Genes that Control Bone and Cartilage Formation 1) wt 2) 3) 4) ko Compare wt vs mutant mouse Identify regulatory networks Identify genes Identify in callus formation disease genes Bioinformatic analysis of genes and functional testing in vitro in vivo chick mouse Test positive candidates in disease models 7 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Mutationen in BMPs verursachen humane Erkrankungen SYM1 SYNS1 GDF5 R438L GDF5 N445T Symphalangismus - Gelenkfusionen BMPs kontrollieren Entwicklung und Aufbau von Knochen Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Therapeutische Anwendung von BMPs 1. Kieferaufbau vor Zahn-Implantation 2. Spinalfusionen bei Bandscheiben-Vorfällen 3. Regeneration bei Knochenbrüchen 8 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Moleküle mit optimierten Eigenschaften: – erhöhte Stabilität – Resistenz gegen Inhibitoren – höhere biologische Aktivität – veränderte Rezeptor-Spezifität Natürlich vorkommende Mutationen identifizieren wichtige funktionelle Domänen und Wirkmechanismen Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies Aktivierende Mutationen • vermehrte Rezeptorbindung micromass analysis • geringere Inhibierbarkeit Mutant B • andere Rezeptoraffinität Mutant A Mutant B 9 Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies GDF5 and its receptor BMPR1B Max Planck Institute for Molecular Genetics Berlin-Brandenburg Center for Regenerative Therapies It looks like a simple process, but of course the science is complex. 10
