Gebala, VM; (2016) Mechanisms of lumen formation during sprouting angiogenesis in vivo. Doctoral thesis , UCL (University College London).

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Abstract

During development, vascular networks expand following a process known as sprouting angiogenesis. New vascular branches arise from pre-existing vessels through the coordinated migration and proliferation of endothelial cells, and eventually connect to form new vascular loops. The functionality of these new vessel segments is dependent on the opening of a central lumen to allow perfusion. While mechanisms of lumen formation during the establishment of the primary vasculature by vasculogenesis have been well characterised, the mechanisms underlying lumen formation during sprouting angiogenesis in vivo are still poorly understood and subject to debate. In this work, I established high spatial and temporal imaging of apical membrane dynamics during sprouting angiogenesis in vivo, both in the zebrafish trunk vasculature and the mouse retina primary inner plexus. By doing so, I identified a novel mechanism of lumen formation, where blood flow expands luminal compartments by driving spherical deformations reminiscent of membrane blebs at the apical membrane of sprouting endothelial cells. Additionally, I show that this process, that I chose to term inverse membrane blebbing, is tightly controlled through local and transient recruitment and contraction of actomyosin at the surface of growing blebs. This mechanism eventually leads to the selective retraction of inverse blebs along the apical membrane, allowing unidirectional lumen expansion. When endothelial cells ability to retract is inhibited through expression of a dominant-negative form of myosin II, cells fail to lumenise or show dilated lumens with multiple unresolved side blebs and branches. Together, this work provides a mechanism for lumen expansion within sprouting endothelial cells in vivo, and identifies for the first time inverse blebbing as a reaction of membranes to high external pressure. In the context of sprouting endothelial cells, I show that a tight equilibrium between the forces exerted by the blood and the contractile responses from the cells is required for lumen expansion within angiogenic sprouts.

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