Author:

Keywords:

sprouting angiogenesis, in vitro model, endothelial invasion, extracellular matrix, collagen, cytoskeleton, mechanobiology, mechanotransduction, cellular forces, pulling forces, computer model, image processing, confocal microscopy, Science & Technology, Life Sciences & Biomedicine, Peripheral Vascular Disease, Cardiovascular System & Cardiology, Sprouting angiogenesis, In vitro model, Endothelial invasion, Extracellular matrix, Collagen, Cytoskeleton, Mechanobiology, Mechanotransduction, Cellular forces, Pulling forces, Computer model, Image processing, Confocal microscopy, CAPILLARY MORPHOGENESIS, MIGRATION, CELLS, QUANTIFICATION, BEHAVIOR, Computer Simulation, Endothelial Cells, Extracellular Matrix, Humans, Models, Cardiovascular, Neovascularization, Physiologic, 1103 Clinical Sciences, 1115 Pharmacology and Pharmaceutical Sciences, Oncology & Carcinogenesis, 3101 Biochemistry and cell biology

Abstract:

Angiogenesis is the formation of new blood vessels from the pre-existing vasculature. It is essential for normal tissue growth and regeneration, but also plays a key role in many diseases. Cytoskeletal components have been shown to be important for angiogenic sprout initiation and maintenance as well as endothelial cell shape control during invasion. The exact nature of cytoskeleton-mediated forces for sprout initiation and progression, however, remains poorly understood. Questions on the importance of tip cell pulling versus stalk cell pushing are to a large extent unanswered, which among others has to do with the difficulty of quantifying and resolving those forces in time and space. We developed methods based on time lapse confocal microscopy and image processing – further termed 4D displacement microscopy - to acquire detailed, spatially and temporally resolved extracellular matrix (ECM) deformations, indicative of cell-ECM mechanical interactions around invading sprouts. We demonstrate that matrix deformations dependent on actin-mediated force generation are spatio-temporally correlated with sprout morphological dynamics. Furthermore, sprout tips were found to exert radially pulling forces on the extracellular matrix, which were quantified by means of a computational model of collagen ECM mechanics. Protrusions from extending sprouts mostly increase their pulling forces, while retracting protrusions mainly reduce their pulling forces. Displacement microscopy analysis further unveiled a characteristic dipole-like deformation pattern along the sprout direction that was consistent among seemingly very different sprout shapes - with oppositely oriented displacements at sprout tip versus sprout base and a transition zone of negligible displacements in between. These results demonstrate that sprout-ECM interactions are dominated by pulling forces and underline the key role of tip cell pulling for sprouting angiogenesis.