93
fibronectin with a distinct structure and organization was uniquely distributed
among other secreted matrix components, such as collagen, tenascin C, versican,
and decorin. Cell matrix adhesions and MT1-MMP activities were reported to orient and localize within this fibrous fibronectin, which is indicative of integrinmediated vascular morphogenesis [190]. In fact, ECs initiate neovascularization by
unfolding soluble fibronectin and depositing a pericellular network of fibrils that
serve as a structural scaffolding on a mechanically ideal substratum for vessel
development [247]. We have studied how such fibronectin organization influences
endothelial tube formation by patterning fibronectin on cell culture surfaces to optimize vasculogenic potential and understand how microstructure influences vascular
tube formation [54]. Alignment of other important ECM proteins, such as collagen,
has also been shown to guide vascular regeneration by enhancing EC organization
and migration [136]. Interestingly, similar effects in vascular organization are
observed when tensile forces are incurred upon vascular fibrin-based constructs,
where vascular network alignment was induced by application of force. Aligned
microvasculature was shown to enhance vascular integration upon implantation in
abdominal muscle [191]. This last study suggests a potential mechanism for organization of ECM components to guide vascular network organization through forceinduced remodeling. It is likely such a mechanism is coupled with ECM degradation
and secretion of new ECM components to establish a microenvironment amenable
to the formation of new blood vessels.
The unique orientation, organization, and nanotopography of fibrous fibronectin
represent features that can be integrated into synthetic scaffolds. Synthetic polymers, like PLGA and polycaprolactone (PCL), can be electrospun to produce various fiber sizes with micro- to nanoscale features that resemble fibrous fibronectin.
We previously showed that surface nanotopography enhanced the formation of
capillary-like structures (CLSs) in vitro [23]. Growing EPCs on grooves that were
600 nm wide reduced their proliferation and enhanced their migration without
changing the expression of EC markers. Moreover, after 6 days of culture, the EPCs
organized into superstructures along the nanogrooves, in significant contrast to the
EPCs grown on planar surfaces (Fig. 4.2b). The addition of Matrigel further induced
the formation of CLSs, with enhanced alignment, organization, and tube length
compared to a flat surface (Fig. 4.2c). This underscores the increasingly important
role of nanotopography in guiding and orienting vascular assembly. When integrated into the tissue-engineered construct—for instance, using filamentous scaffold geometry [75] and micropatterning [55, 108, 165]—the orientation and
structure of the engineered vasculature can be controlled.
Regulating Matrix Mechanics
It has become increasingly evident that the biomechanical properties of the ECM,
such as matrix orientation and mechanics, profoundly influence the control of vascular morphogenesis. Due to their versatility with respect to mechanical properties
(e.g., cross-linking density, pore sizes, and topography), synthetic biomaterials have
4 Hypoxia and Matrix Manipulation for Vascular Engineering
fibronectin with a distinct structure and organization was uniquely distributed
among other secreted matrix components, such as collagen, tenascin C, versican,
and decorin. Cell matrix adhesions and MT1-MMP activities were reported to orient and localize within this fibrous fibronectin, which is indicative of integrinmediated vascular morphogenesis [190]. In fact, ECs initiate neovascularization by
unfolding soluble fibronectin and depositing a pericellular network of fibrils that
serve as a structural scaffolding on a mechanically ideal substratum for vessel
development [247]. We have studied how such fibronectin organization influences
endothelial tube formation by patterning fibronectin on cell culture surfaces to optimize vasculogenic potential and understand how microstructure influences vascular
tube formation [54]. Alignment of other important ECM proteins, such as collagen,
has also been shown to guide vascular regeneration by enhancing EC organization
and migration [136]. Interestingly, similar effects in vascular organization are
observed when tensile forces are incurred upon vascular fibrin-based constructs,
where vascular network alignment was induced by application of force. Aligned
microvasculature was shown to enhance vascular integration upon implantation in
abdominal muscle [191]. This last study suggests a potential mechanism for organization of ECM components to guide vascular network organization through forceinduced remodeling. It is likely such a mechanism is coupled with ECM degradation
and secretion of new ECM components to establish a microenvironment amenable
to the formation of new blood vessels.
The unique orientation, organization, and nanotopography of fibrous fibronectin
represent features that can be integrated into synthetic scaffolds. Synthetic polymers, like PLGA and polycaprolactone (PCL), can be electrospun to produce various fiber sizes with micro- to nanoscale features that resemble fibrous fibronectin.
We previously showed that surface nanotopography enhanced the formation of
capillary-like structures (CLSs) in vitro [23]. Growing EPCs on grooves that were
600 nm wide reduced their proliferation and enhanced their migration without
changing the expression of EC markers. Moreover, after 6 days of culture, the EPCs
organized into superstructures along the nanogrooves, in significant contrast to the
EPCs grown on planar surfaces (Fig. 4.2b). The addition of Matrigel further induced
the formation of CLSs, with enhanced alignment, organization, and tube length
compared to a flat surface (Fig. 4.2c). This underscores the increasingly important
role of nanotopography in guiding and orienting vascular assembly. When integrated into the tissue-engineered construct—for instance, using filamentous scaffold geometry [75] and micropatterning [55, 108, 165]—the orientation and
structure of the engineered vasculature can be controlled.
Regulating Matrix Mechanics
It has become increasingly evident that the biomechanical properties of the ECM,
such as matrix orientation and mechanics, profoundly influence the control of vascular morphogenesis. Due to their versatility with respect to mechanical properties
(e.g., cross-linking density, pore sizes, and topography), synthetic biomaterials have
4 Hypoxia and Matrix Manipulation for Vascular Engineering