Topography-driven alterations in endothelial cell phenotype and contact guidance

Abstract

Understanding how endothelial cell phenotype is affected by topography could improve the design of new tools for tissue engineering as many tissue engineering approaches make use of topography-mediated cell stimulation. Therefore, we cultured human pulmonary microvascular endothelial cells (ECs) on a directional topographical gradient to screen the EC vascular-like network formation and alignment response to nano to microsized topographies. The cell response was evaluated by microscopy. We found that ECs formed unstable vascular-like networks that aggregated in the smaller topographies and flat parts whereas ECs themselves aligned on the larger topographies. Subsequently, we designed a mixed topography where we could explore the network formation and proliferative properties of these ECs by live imaging for three days. Vascular-like network formation continued to be unstable on the topography and were only produced on the flat areas and a fibronectin coating did not improve the network stability. However, an instructive adipose tissue-derived stromal cell (ASC) coating provided the correct environment to sustain the vascular-like networks, which were still affected by the topography underneath. It was concluded that large microsized topographies inhibit vascular endothelial network formation but not proliferation and flat and nano/microsized topographies allow formation of early networks that can be stabilized by using an ASC instructive layer.

Topography-Mediated Myotube and Endothelial Alignment, Differentiation, and Extracellular Matrix Organization for Skeletal Muscle Engineering

“…Directional topography caused cells to align ( Figure 3 c). After five days on flat PDMS, ECs had detached and formed aggregates, whereas the ECs on the directional topography remain aligned ( Figure 3 d), which corroborates our earlier findings on mixed topographies that ECs tend to detach and aggregate [ 17 ].…”

“…Directional topography caused cells to align (Figure 3c). After five days on flat PDMS, ECs had detached and formed aggregates, whereas the ECs on the directional topography remain aligned (Figure 3d), which corroborates our earlier findings on mixed topographies that ECs tend to detach and aggregate [17]. After two days of co-culture, endothelial cells readily attached and followed the myotubes directionality irrespective of the underlying substrate (Figure 4a).…”

“…Directional topography caused cells to align (Figure 3c). After five days on flat PDMS, ECs had detached and formed aggregates, whereas the ECs on the directional topography remain aligned (Figure 3d), which corroborates our earlier findings on mixed topographies that ECs tend to detach and aggregate [17]. Myotubes and ECs attached to the directional topography ( Figure 3) while adhering poorly to flat PDMS and readily detached within five days, which was only marginally improved by prior coating with gelatin [12].…”

“…In our system, the lack of deposition of adequate amounts of fibronectin may have hampered the development of adhered ECs to mature networks. Of note, this is not a limitation of the use of HPMECs, because the topographical substrates by themselves warranted spontaneous network formation [17]. Our data appear to conflict with those of Nagamori et al who reported that embryonic endothelial cells (HUVEC) readily formed networks on myoblast sheets produced by detachment from thermoresponsive flat substrates [38].…”

“…The interaction between human myoblasts and endothelial cells in an aligned topography is pertinent for more sophisticated and functional skeletal muscle engineering but has not yet been adequately studied [14][15][16]. We recently published that also vascular endothelial cells prefer linear topographies for proliferation over flat surfaces while network formation did not follow linear topography [17]. This was expected, because networks are interconnected endothelial with a honeycomb appearance in vitro as well as in vivo.…”