Junctional remodeling during angiogenesis : the role of Claudin5 during ISV formation in Zebrafish

During sprouting angiogenesis several cellular activities are required such as cell shape changes, cell migration, cell proliferation and cell rearrangement. These processes have to be tightly regulated in order to allow cellular dynamics on one side and maintain tissue integrity on the other side. Cell-cell contacts are the main mediators of dynamic cell behaviors during morphogenesis. These contacts are composed of adherens and tight junctions.
To better understand the role of tight junctions during the morphogenetic events of angiogenesis, I have analyzed the function of the endothelial specific Cldn5b during the process of intersegmental vessel (ISV) formation. Although MO-mediated knock-down of cldn5b does not alter the initial outgrowth of endothelial cells (ECs) from the dorsal aorta (DA), it does lead to a halt of EC migration at the level of the horizontal myoseptum (HMS). Furthermore, neighboring ISVs do not connect within the dorsal-longitudinal-anastomotic vessel (DLAV) and detach from the DA. In contrast, the adherens junction specific protein VE-cadherin rather exerts a cell rearrangement function. The different expression pattern of Cldn5b and VE-cadherin support these finding. While VE-cadherin is expressed in every EC of the ISV, Cldn5b is temporally downregulated in early sprouting ECs. Furthermore, a clear reduction of cell number in the ISVs was observed in cldn5b morphants. The latter observation, together with a possible reduction in adhesion between ECs are suggested to cause the phenotypes observed in cldn5b knock-down embryos. Together, this study suggests for the first time a possible role of a Cldn during a morphogenetic event.
Additionally, I generated mutant fish lacking cldn5b gene function by using Zinc-Finger Nucleases. The analysis of these mutant fish will help to understand the functional role of a core component of tight junctions during vertebrate blood vessel development. However, the preliminary analysis show only a very weak ISV phenotype compared to cldn5b morphants. I will discuss several ideas explaining the discrepancy between cldn5b morphants and mutants.