Matrix Mechanics and Cell Contractility in Angiogenesis - Mechanical and Chemical Signaling in Angiogenesis - page 144

2.2 Endothelial Cell Shape and Cytoskeletal Tension
Regulate Proliferation
Endothelial cell growth is critical to angiogenesis, and shape has emerged as an
important regulator of proliferation. Early experiments with ECs determined that
increases in spread area due to increasing density of substrate-bound fibronectin
are accompanied by an increase in cell proliferation [ 31 ]. The investigation of cell
shape on the control of cell cycle progression indicated that ECs must remain
spread for most of G 1 to enter S phase [ 32 ]. As ECs spread during G 1 , increasing
nuclear volume and DNA synthesis predict the proliferation response [ 33 ].
The use of micropatterned substrates that control cell spreading has identified
cell shape as a switch between EC life and death [ 34 , 35 ]. ECs grown on
microadhesive islands less than 500 lm 2 undergo apoptosis, but on substrates
greater than 1,500 lm 2 cells spread and grow [ 36 ]. When grown on small adhesive
islands that prevent spreading, cells fail to progress from G 1 to S phase despite
normal activation of the Erk1/2 MAPK pathway [ 37 ]. Failure to progress into S
phase is attributed to an inability to increase cyclin D, reduce the cyclin-dependent
kinase inhibitor p27 Kip1 , and phosphorylate Rb in late G 1 (Fig. 2 ). Changes in EC
shape and spreading that predict growth are tied to tension in the cytoskeleton, and
cell cycle progression can be blocked by cytoskeletal disruption. Disruption of the
actin filament network with cytochalasin D or microtubules with nocodazole leads
to apoptosis similar to that seen in cells on small adhesive islands that restrict
spreading [ 38 ]. Simultaneous disruption of actin and microtubules leads to
increased rounding, dephosphorylation of the serine/threonine kinase Akt (also
known as Protein Kinase B), decreased expression of Bcl-2, and increased caspase
activity, hallmarks of apoptosis (Fig. 2 ).
Cytoskeletal tension and cell spreading have been identified as a late G 1
restriction point that acts independently of MAPK/Erk signaling to promote the
G 1 -S transition during growth [ 39 ]. When actin polymerization is disrupted with
cytochalasin D, cyclin D expression is decreased, ECs accumulate p27 Kip1 , and
cell cycle progression is arrested. Separate work determined that when EC
spreading is inhibited by using substrates with reduced ligand density, cells also
accumulate p27 Kip1 and arrest in mid G 1 [ 40 ]. This response is rescued with
constitutively active Rho that increases Skp2 expression (required for degradation
of p27 Kip1 and progression through G 1 ) and promotes G 1 progression by altering
the balance of mDia1 and ROCK, effectors of Rho that mediate cell contractility
and actin cytoskeletal organization (Fig. 2 ). These findings indicate that EC
cytoskeletal tension and shape are important cell cycle checkpoints that mediate
cell cycle progression.
Cell-cell interactions also contribute to EC growth decisions by modulating
cytoskeletal tension. VE-cadherin-mediated cell-cell contacts decrease cell
spreading and proliferation, but if spreading is prevented by growing cells on
micropatterned substrates, proliferation increases via Rho signaling and cytoskeletal
tension [ 41 ]. Use of dominant negative RhoA can block cell-cell contact-mediated