German Center for Cardiovascular Research (DZHK)

6 ARTICLES PUBLISHED IN JoVE

Immunology and Infection

Engineering and Evolution of Synthetic Adeno-Associated Virus (AAV) Gene Therapy Vectors via DNA Family Shuffling

Eike Kienle^*1, Elena Senís^*1, Kathleen Börner², Dominik Niopek¹, Ellen Wiedtke¹, Stefanie Grosse¹, Dirk Grimm¹

¹Cluster of Excellence CellNetworks, Department of Infectious Diseases, Virology, Heidelberg University, ²Department of Infectious Diseases, Virology, Heidelberg University

We demonstrate the basic technique to molecularly engineer and evolve synthetic Adeno-associated viral (AAV) gene therapy vectors via DNA family shuffling. Moreover, we provide general guidelines and representative examples for selection and analysis of individual chimeric capsids with enhanced properties on target cells in culture or in mice.

Developmental Biology

Processing of Human Cardiac Tissue Toward Extracellular Matrix Self-assembling Hydrogel for In Vitro and In Vivo Applications

Matthias Becker^1,2, Janita A. Maring^1,2, Barbara Oberwallner², Benjamin Kappler², Oliver Klein^1,2, Volkmar Falk^2,3,4,5, Christof Stamm^2,3,4

¹Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, ²Berlin-Brandenburg Center for Regenerative Therapies (BCRT), ³German Center for Cardiovascular Research (DZHK), ⁴Deutsches Herzzentrum Berlin (DHZB), ⁵Department of Cardiovascular Surgery, Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health

This protocol describes the complete decellularization of human myocardium while preserving its extracellular matrix components. Further processing of the extracellular matrix results in the production of microparticles and a cytoprotective self-assembling hydrogel.

Immunology and Infection

Visualizing Leukocyte Rolling and Adhesion in Angiotensin II-Infused Mice: Techniques and Pitfalls

Jeremy Lagrange¹, Sabine Kossmann^1,2, Klytaimnistra Kiouptsi¹, Philip Wenzel^1,2,3

¹Center for Thrombosis and Hemostasis Mainz, University Medical Center Mainz, ²Center for Cardiology, University Medical Center Mainz, ³German Center for Cardiovascular Research (DZHK)

This manuscript describes the use of transgenic reporter mice and different administration routes of fluorescent dyes in angiotensin II-induced hypertension using intravital video microscopy of blood vessels to evaluate the activation of immune cells and their ability to roll and adhere to the endothelium.

Medicine

Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction

David Bode^1,2, Tim Guthof¹, Burkert M. Pieske^1,2, Frank R. Heinzel^1,2, Felix Hohendanner^1,2

¹Department of Internal Medicine and Cardiology, Charité University Medicine, ²German Center for Cardiovascular Research (DZHK)

Here, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction. A manual regulation of intraluminal pressure of cardiac cavities is implemented to yield functionally intact myocytes suitable for excitation-contraction-coupling studies.

Neuroscience

Analysis of Cerebral Vasospasm in a Murine Model of Subarachnoid Hemorrhage with High Frequency Transcranial Duplex Ultrasound

Axel Neulen^*1, Michael Molitor^*2,3,4, Michael Kosterhon^*1, Tobias Pantel¹, Susanne H. Karbach^2,3,4, Philip Wenzel^2,3,4, Thomas Gaul⁵, Florian Ringel¹, Serge C. Thal⁵

¹Department of Neurosurgery, University Medical Center of the Johannes Gutenberg-University of Mainz, ²Center for Cardiology - Cardiology I, University Medical Center of the Johannes Gutenberg-University of Mainz, ³Center for Thrombosis and Hemostasis (CTH), University Medical Center of the Johannes Gutenberg-University of Mainz, ⁴German Center for Cardiovascular Research (DZHK) - Partner site Rhine-Main, ⁵Department of Anesthesiology, University Medical Center of the Johannes Gutenberg-University of Mainz

The aim of this manuscript is to present a sonography-based method that allows in vivo imaging of blood flow in cerebral arteries in mice. We demonstrate its application to determine changes in blood flow velocities associated with vasospasm in murine models of subarachnoid hemorrhage (SAH).

Bioengineering

Isolation of Next-Generation Gene Therapy Vectors through Engineering, Barcoding, and Screening of Adeno-Associated Virus (AAV) Capsid Variants

Kleopatra Rapti^1,2, Olena Maiakovska^1,2, Jonas Becker^1,2, Joanna Szumska^1,2, Margarita Zayas^1,2, Felix Bubeck^1,2, Jixin Liu^1,2, Emma Gerstmann^1,2, Chiara Krämer^1,2, Ellen Wiedtke^1,2, Dirk Grimm^1,2,3,4,5

¹Department of Infectious Diseases/Virology, Section Viral Vector Technologies, Medical Faculty, University of Heidelberg, ²BioQuant Center, BQ0030, University of Heidelberg, ³Cluster of Excellence CellNetworks, ⁴German Center for Infection Research (DZIF), ⁵German Center for Cardiovascular Research (DZHK)

AAV peptide display library generation and subsequent validation through the barcoding of candidates with novel properties for the creation of next-generation AAVs.