Matrix Mechanics and Cell Contractility in Angiogenesis - Mechanical and Chemical Signaling in Angiogenesis - page 143

cell-cell interactions. Increasing RhoA-mediated myosin-generated contraction
(e.g. treatment with thrombin) disrupts cell-cell contacts and adherens junction
assembly [ 21 ]. This response is sensitive to the stiffness of the mechanical
microenvironment. Increased matrix stiffness promotes disrupted VE-cadherin
mediated cell-cell junctions and results in an increase in EC monolayer perme-
ability [ 22 ]. These data are indicative of crosstalk and feedback mechanisms that
exist between integrins and cadherins [ 23 ] that mediate cellular force balance, and
suggest

that matrix stiffness alters endothelial cell-cell and cell-matrix

connectivity.
In addition to mediating changes in cell spreading, matrix stiffness may directly
impact traction force generation. As substrate stiffness is increased, there is an
increase in endothelial cell traction force generation [ 13 ] (Fig. 1 c). Measurements
of force generation and cell area coupled with regression modeling indicate that
both area and matrix stiffness are significant predictors of EC traction forces in
single cells and cells in contact [ 13 ]. While the total magnitude of the force exerted
by ECs is linearly related to cell area during spreading events, cells are capable of
exerting significant traction forces in the absence of notable focal adhesions or
stress fiber formation [ 12 ], typical attributes of well-spread cells. These results
indicate that matrix stiffness alters EC traction force generation, and suggest that
ECs may utilize small nascent focal complexes to exert traction forces, a mech-
anism first described in fibroblasts [ 24 ].
Changes in matrix stiffness-mediated EC contractility associated with spreading
or focal adhesion organization may be due to cell shape changes that physically
alter the localization and activity of contractile adaptor proteins. During cell
spreading after plating, the actin-binding protein filamin binds the Rho GTPase
activating protein (GAP) p190 (p190RhoGAP) thus preventing its accumulation in
membrane lipid rafts and allowing high Rho activity [ 25 ]. When cells are spread
the protease calpain cleaves filamin allowing the accumulation of p190RhoGAP in
lipid rafts that inhibit Rho activity. These data indicate that cell spreading alters
the localization and activity of mediators of cellular contractility. They suggest
that matrix stiffness alters cell shape and contractility via changes in protein
localization; however, more work is needed to understand the crosstalk of intra-
cellular proteins that participate in these mechanisms.
Notably, p190RhoGAP has been shown to regulate capillary formation in vitro
and retinal angiogenesis in vivo by altering the transcription factors TFII-I and
GATA2, regulators of VEGFR2 expression [ 26 ]. Moreover, GATA2 and VEGFR2
expression levels increase with increasing substrate stiffness. Interestingly, VEGF
stimulates stress fiber and focal adhesion organization [ 27 ] through Rho and
ROCK signaling [ 28 ]. More recent work indicates that VEGF induces a two-fold
contractile response mediated by VEGFR2 and ROCK [ 29 ]. These data suggest
that angiogenesis is mediated by EC sensitivity to growth factor signaling that is
intimately associated with matrix stiffness and the contractile machinery of the
cell.