University of Rostock View Institution's Website 14 articles published in JoVE Cancer Research Creation and Maintenance of a Living Biobank - How We Do It Florian Bürtin1, Stephanie Matschos2, Friedrich Prall3, Christina S. Mullins2, Mathias Krohn2, Michael Linnebacher2 1Department of General, Visceral, Vascular and Transplantation Surgery, University Medical Center Rostock, University of Rostock, 2Molecular Oncology and Immunotherapy, Department of General, Visceral, Vascular and Transplantation Surgery, University Medical Center Rostock, 3Institute of Pathology, University Medical Center Rostock In the following work, we describe the consecutive steps necessary for the establishment of a large biobank of colorectal and pancreatic cancer. Biology Analyzing the α-Actinin Network in Human iPSC-Derived Cardiomyocytes Using Single Molecule Localization Microscopy Lisa Johann1,2, Oleksandra Chabanovska1,2, Cajetan Immanuel Lang3, Robert David1,2, Heiko Lemcke1,2 1Department of Cardiac Surgery, Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Rostock University Medical Center, 2Faculty of Interdisciplinary Research, Department Life, Light & Matter, University Rostock, 3Department of Cardiology, Rostock University Medical Center The formation of a proper sarcomere network is important for the maturation of iPSC-derived cardiomyocytes. We present a super resolution-based approach that allows for the quantitative evaluation of the structural maturation of stem cell derived cardiomyocytes, to improve culture conditions promoting cardiac development. Medicine Intrathecal Application of a Fluorescent Dye for the Identification of Cerebrospinal Fluid Leaks in Cochlear Malformation Nora M. Weiss1, Ingo Andus2, Armin Schneider3, Sönke Langner4, Stefanie Schröder1, Sebastian P. Schraven1, Robert Mlynski1 1Department of Otorhinolaryngology, Head and Neck Surgery, University Medical Center Rostock, 2Rostock University, 3ARRI Medical GmbH, 4Department of Radiology and Institute of Diagnostic and Interventional Radiology, University Medical Center Rostock Intrathecally applied fluorescein is used to achieve intraoperative visualization of CSF leaks. This protocol describes a lumbar puncture, the application of 5% fluorescein, and intraoperative visualization using a fully digital microscope. Medicine Synergizing Antegrade Endoscopic with Bridging Vein Harvesting for Improvement of Great Saphenous Vein Graft Quality from the Lower Leg Christian Klopsch1, Alexander Kaminski1, Friedrich Prall2, Pascal Dohmen1,3 1Department of Cardiac Surgery, Heart Center Rostock, Rostock University Medical Center, University of Rostock, 2Institute of Pathology, Rostock University Medical Center, University of Rostock, 3Department of Cardiothoracic Surgery, Faculty of Health Science, University of the Free State Presented here is a protocol for antegrade endoscopic vein harvesting from the lower leg, which can safely be introduced in routine coronary artery bypass grafting. Vein grafts present excellent graft quality following this standardized protocol with positioning of the legs, minimally invasive access to the vein, and antegrade endoscopic vein harvesting. Chemistry Extraction of Lignin with High β-O-4 Content by Mild Ethanol Extraction and Its Effect on the Depolymerization Yield Douwe S. Zijlstra1, Alessandra de Santi2, Bert Oldenburger1,2, Johannes de Vries2,3, Katalin Barta2, Peter J. Deuss1 1Department of Chemical Engineering (ENTEG), University of Groningen, 2Stratingh Institute for Chemistry, University of Groningen, 3Leibniz-Insitut f ür Katalyse e.V., Universität Rostock Here, we present a protocol to perform ethanol extraction of lignin from several biomass sources. The effect of the extraction conditions on the lignin yield and β-O-4 content are presented. Selective depolymerization is performed on the obtained lignins to obtain high aromatic monomer products. Genetics Isolation, Characterization and MicroRNA-based Genetic Modification of Human Dental Follicle Stem Cells Paula Müller*1,2, Katharina Ekat*3, Anne Brosemann3, Anne Köntges3, Robert David1,2, Hermann Lang3 1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, Rostock University Medical Center, 2Department Life, Light and Matter of the Interdisciplinary Faculty at Rostock University, 3Department of Operative Dentistry and Periodontology, Rostock University Medical Center This protocol describes the transient genetic engineering of dental stem cells extracted from the human dental follicle. The applied non-viral modification strategy may become a basis for the improvement of therapeutic stem cell products. Bioengineering Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting Frauke Hausburg*1,2, Paula Müller*1,2, Natalia Voronina*1, Gustav Steinhoff1,2, Robert David1,2 1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, Rostock University Medical Center, 2Department Life, Light and Matter of the Interdisciplinary Faculty, Rostock University This protocol illustrates a safe and efficient procedure to modify CD133+ hematopoietic stem cells. The presented non-viral, magnetic polyplex-based approach may provide a basis for the optimization of therapeutic stem cell effects as well as for monitoring the administered cell product via magnetic resonance imaging. Biology Analysis of the Gap Junction-dependent Transfer of miRNA with 3D-FRAP Microscopy Heiko Lemcke1,2,3, Natalia Voronina1,2,3, Gustav Steinhoff1,2,3, Robert David1,2,3 1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), 2Department of Cardiac Surgery, University of Rostock, 3Department of Life, Light and Matter of the Interdisciplinary Faculty, University of Rostock Here, we describe the application of three-dimensional fluorescence recovery after photobleaching (3D-FRAP) for the analysis of the gap junction-dependent shuttling of miRNA. In contrast to commonly applied methods, 3D-FRAP allows for the quantification of the intercellular transfer of small RNAs in real time, with high spatio-temporal resolution. Medicine Preparation and In Vitro Characterization of Magnetized miR-modified Endothelial Cells Natalia Voronina1, Heiko Lemcke1, Frank Wiekhorst2, Jens-Peter Kühn3, Markus Frank4, Gustav Steinhoff1, Robert David1 1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, University of Rostock, 2Physikalisch-Technische Bundesanstalt, 3Department of Radiology and Neuroradiology, Ernst-Moritz-Arndt-University Greifswald, 4Electron Microscopy Center, University of Rostock This manuscript describes the efficient, non-viral delivery of miR to endothelial cells by a PEI/MNP vector and their magnetization. Thus, in addition to genetic modification, this approach allows for magnetic cell guidance and MRI detectability. The technique can be used to improve the characteristics of therapeutic cell products. Developmental Biology Generation of Murine Cardiac Pacemaker Cell Aggregates Based on ES-Cell-Programming in Combination with Myh6-Promoter-Selection Christian Rimmbach1, Julia J. Jung1, Robert David1 1Reference and Translation Center for Cardiac Stem Cell Therapy, University of Rostock This protocol describes how to produce functional sinus nodal tissue from murine pluripotent stem cells (PSC). T-Box3 (TBX3) overexpression plus cardiac Myosin-heavy-chain (Myh6) promoter antibiotic selection leads to highly pure pacemaker cell aggregates. These “Induced-sinoatrial-bodies” (“iSABs”) contain over 80% pacemaker cells, show highly increased beating rates and are able to pace myocardium ex vivo. Biology Intravital Microscopy of the Microcirculation in the Mouse Cremaster Muscle for the Analysis of Peripheral Stem Cell Migration Peter Donndorf1, Marion Ludwig1, Fabian Wildschütz1, Dritan Useini1, Alexander Kaminski1, Brigitte Vollmar2, Gustav Steinhoff1 1Reference and Translation Centre for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, University Rostock, 2Institute for Experimental Surgery, University of Rostock Intravital microscopy of the mouse M. cremaster microcirculation offers a unique and well-standardized in vivo model for the analysis of peripheral bone marrow stem cell migration. Biology Gene Transfer into Older Chicken Embryos by ex ovo Electroporation Jiankai Luo1, Xin Yan1, Juntang Lin2, Arndt Rolfs1 1Albrecht-Kossel-Institute for Neuroregeneration, School of Medicine University of Rostock, 2Institute of Anatomy I, School of Medicine University of Jena A method of gene transfer into chicken embryos at later incubation stages (older than Hamburger and Hamilton stage (HH) 22) is described. This method overcomes disadvantages of in ovo electroporation applied to older chicken embryos and is a useful technique to study gene function and regulation at older developmental stages. Medicine A New Single Chamber Implantable Defibrillator with Atrial Sensing: A Practical Demonstration of Sensing and Ease of Implantation Dietmar Bänsch1, Ralph Schneider1, Ibrahim Akin1, Cristoph A. Nienaber1 1Heart Center Rostock, University Hospital of Rostock, Germany Dual-chamber implantable cardioverter-defibrillators (ICDs) may improve detection of atrial fibrillation as well as differentiation of tachycardias. However, this advantage is undermined by complications associated with the second electrode, which is required in conventional dual chamber devices. Therefore, BIOTRONIK has developed a new electrode called the LinoxSMART S DX that, when used in conjunction with the Lumax DX ICD, offers dual-chamber detection without the risks associated with the second electrode. Bioengineering Cultivation of Human Neural Progenitor Cells in a 3-dimensional Self-assembling Peptide Hydrogel Andrea Liedmann1, Arndt Rolfs1, Moritz J. Frech1 1Albrecht-Kossel-Institute for Neuroregeneration, University of Rostock Here we describe the use of a self-assembling 3-dimensional scaffold to culture human neural progenitor cells. We present a protocol to release the cells from the scaffolds to be analysed subsequently e.g. by flow cytometry. This protocol might be adapted to other cell types to perform detailed mechanistically studies.