3D PRINTING AND PATTERNING VASCULATURE IN ENGINEERED TISSUES - 3D Bioprinting and Nanotechnology in Tissue Engineering and Regenerative Medicine - page 169

is known to be extremely sensitive to small perturbations in fluid dynamics and microenvironment,
rapidly leading to clotting ( McGuigan and Sefton, 2007 ). Here we discuss approaches to pattern per-
fusable channels and networks in engineered tissues, motivated by limitations in controlling individual
pore size, shape, arrangement, and interconnectivity of microdevices.
Photolithography can typically pattern only a single layer of material at a time. Thus, strategies
combining photolithographic processes with layer-by-layer lamination methods represent an impor-
tant approach to achieve 3D scaffolds with high-resolution microstructures. To this end, Bettinger
et al . developed a vasculature construct out of poly(glycerol-sebacate) (PGS), a tough, biodegradable
elastomer with superior mechanical properties ( Bettinger et al., 2005 ). Multiple layers of PGS vascu-
lature construct were obtained by stacking and aligning layers, followed by a thermal bonding process.
Although the authors demonstrated that cells seeded on the fabricated scaffold resulted in normal meta-
bolic activity, the ability to encapsulate cells in these constructs was limited by the harsh fabrication
process. Furthermore, the ability to precisely form microstructures with encapsulated cells in complex
3D structures cannot easily be obtained using this approach.
King et al. developed a novel technique for fabrication of high-resolution, high-precision features
by combining a thermal fusion bonding process after fabrication of a single layer of microfluidic
network composed of thermoplastic PLGA ( King et al., 2004 ). This optimized approach involves
conventional photolithography and silicon micromachining of single layers, followed by precise
alignment and stacking of interconnected micropatterned films and thermal sealing, to obtain highly
branched, multilayer PLGA microfluidic networks. Moreover, the authors were able to demonstrate
the formation of 2 m m wide channels, close to the physical limits of traditional photolithography,
all the way up to channels in the millimeter range. When perfused, single-layer networks showed
no sign of leaks, occlusions, or channel-to-channel cross-talk, and demonstrated linear pressure-
flow characteristics. Lactide has also been incorporated into a copolymer with PEG to form micro-
channels by selective degradation of bulk photopolymerized hydrogel substrates ( Chiu et al., 2009 ).
While the bulk of the scaffold was made from poly(ethylene glycol) diacrylate (PEGDA) by expos-
ing light without a photomask, a photomask was used when PEG-PLLA-DA was injected into the
system to create smaller patterns on top of the previous layer. Then, the spaces between the patterned
strips were injected with PEGDA and cured. Channels were obtained in scaffolds by repeating the
steps to create multilayered constructs and then incubating the construct in high pH environment to
accelerate degradation of the PEG-PLLA-DA copolymer. This route for microchannel fabrication
offers a 3D architecture with interconnected microchannels that can be obtained from biocompatible
materials.
Inspired by the complex, multiscale microvascular network of leaf venation, He et al. fabri-
cated nature-inspired microfluidic networks for perfusable tissue constructs ( He et al., 2013 ). Their
technique involved a microreplication method to mimic the microvascular network of natural leaf
in synthetic substrates by digesting leafs to expose venation networks, sputtering with a thin layer
of chrome, and then using the chrome-coated leaf venation as photomask to obtain a silicon mold
with leaf venation to fabricate biomaterial hydrogels with multiscale vascular channels ranging
from 30 m m to 1 mm in diameter. Seeding cells in this system resulted in high viability of endo-
thelial cells, proliferation, and spreading along the microfluidic channels during the short culture
period. The channels were perfused by mounting the hydrogels on an incline plate and introducing
fluid from an inlet in a drop-wise manner, whose flow was induced by gravity. Further work by
Wu et al. studied the transport efficiency of microvascular networks inspired by the hierarchical,