Objectives

The main objective of this Workpackage is the Investigation of radiation effects on cardiac cell integrity in CVD by studying morphological and functional properties of cardiomyocytes and endothelial cells in vitro.

Task 1:  Study of morphological and functional properties of cardiomyocytes
Task 2:  Investigation of endothelial interactions

Workpackage description

• This Workpackage will isolate and culture cardiomyocytes at different times after irradiation from irradiated hearts. Samples of these cultures will be sent to the involved partners for co-culture studies and for proteomics studies.
• The activation of the Rho/ROCK pathway will be studied. The organization of actin-based cytoskeletal structures and regulation of actin-myosin filament bundles that control cell contractility will be investigated using pharmacological inhibitors like statins and genetic knocking down approaches.
• Endothelial cells will be isolated at different times after radiation and cultivated further. Afterwards changes changes in permeability of cultured endothelial cells monolayers, and changes in endothelial cell cytoskeleton (e.g. actin) and junction (e.g. VE-cadherin) will be analysed as function of radiation dose and time. 
• Additionally, long-term changes of intercellular communication in endothelial cultures will be studied by local irradiation of endothelial cells using X-ray microbeam approaches and testing for functional changes in permeability of monolayer sheets and differences in inter- and intracellular signalling mechanisms. 
• The angiogenic properties of endothelial cells will be investigated  by studying the organisation of cells into 3D capillary-like structures using in vitro models (also in co-culture with cardiomyocytes) and by evaluating different mechanisms of endothelial cell migration, namely chemotaxis, haptotaxis and mechanotaxis. Emphasis will be placed on studying signalling pathways that converge to cytoskeletal remodelling, and which are required for 3D morphogenesis and migration. The contribution of stress stimuli and hypoxia or hypoxia-reoxygenation on endothelial angiogenic properties will also be investigated. Finally, long-term changes in protein patterns will be investigated using proteomics approaches.

WP leader

Dr. Chryso Kanthou (USFD), Senior Lecturer within the TMG, is a Cell and Molecular Vascular Biologist with expertise in both the cancer and cardiovascular fields. She has a substantial record in the successful isolation and culture of primary vascular-derived cells (smooth muscle and endothelial cells). She also has extensive experience in analysing at the cellular and molecular level, angiogenic and other functional cell responses, including responses to radiation, using in vitro models of endothelial cells.

Participating consortium partners

Technische Universtität München, Germany
Institut de Radioprotection et de Sureté nucléaire Villejuif, France
University of Belfast, United Kingdom
Maria Sklodowska - Curie Cancer Institute, Poland