Models for angiogenesis on micro-structured surfaces

Endothelial cell (EC) migration is an essential process in
angiogenesis as ECs sprout from preexisting vessels, following
chemotactic gradients. However, most of the data obtained about EC
migration has been acquired in artificial two dimensional (2D) cell
culture environments. Recent reports showed that migration in
fibrillary environments can be mimicked by spatial confinement,
achieved by micro patterning techniques (Doyle et al. 2009). In the
first part of this work it was investigated whether a model system
based on linearly structured surfaces allows to draw conclusions
about the migration of ECs in fibrillary 3D collagen matrices. In
order to estimate the cellular behavior of ECs on linearly
structured surfaces, a comprehensive cell biological analysis was
performed. ECs on narrow 3 µm wide tracks (also termed 1D in the
following) migrated less efficient in comparison to ECs on broader
tracks in regard to mean velocity, persistence, and run velocity.
Additionally, ECs in 1D displayed a distinct actin cytoskeleton
architecture, compressed nuclei, and different orientation of the
centrosome in comparison to ECs on wider tracks. The frequent
directional changes of ECs on narrow tracks were accompanied by
pronounced membrane blebbing, while migrating and elongated cells
displayed a lamellipodium as cellular protrusion. This behavior was
contractility-dependent as both modes were provoked by using
Blebbistatin or Calyculin A, respectively. The comparison between
1D and 3D migrating cells revealed a striking similarity in actin
cytoskeleton architecture and in switching between two
morphological modes. Cells migrating in 3D moved slower but more
persistent after Blebbistatin treatment, which was likewise the
case for cells migrating in 1D. In contrast to this, cells in the
2D system migrated faster but less persistent after Blebbistatin
treatment. A Rac1 inhibitor used in this study showed the tendency
to influence the migratory potential similarly in 1D and 3D, in
contrast to 2D. However, a microtubule disrupting agent displayed
different effects in 1D and 3D. These experiments demonstrated that
the 1D system allows to draw conclusions about certain aspects of
3D migration. Thus, using this 1D migration system, important
aspects of 3D migration can be mimicked in a highly controlled
setting. In the second part of this work, a system for artificial
tip cell formation was investigated. For the analysis of tip and
stalk cells specifically structured surfaces were designed. These
structures provided areas allowing only a restricted number of
cell-cell contacts and areas allowing a high number of cell-cell
contacts. ECs with a low number of cell-cell contacts displayed
increased VEGFR2 expression levels in comparison to cells with a
high number of cell-cell contacts, a phenomenon which was inhibited
by using a Notch signaling inhibitor. This system will be a useful
tool in the future to decipher tip and stalk cell competition
within a defined cellular population and a defined microscopic
frame