CRANIOFACIAL BONE - 3D Bioprinting and Nanotechnology in Tissue Engineering and Regenerative Medicine - page 219

bonding, melt molding, and gas foaming), only a few lead to the formation of 3D structures with na-
noscale features to mimic the natural bone ECM structure. These include production of nanofibers by
various techniques including electrospinning, self-assembly, and thermally induced phase separation.
Electrospinning is based on production of fibers on a grounded collector from a polymer solution by the
application of an electric field and can be applied with both natural (e.g. fibrinogen on a thrombin collec-
tion bath to form electrospun fibrin fibers) ( Zhang et al., 2014 ) and synthetic such as poly(ethersulfone)
( Ren et al., 2013 )) polymers to produce scaffolds with nano- and microfibers. Thin fiber layers (mats)
obtained through electrospinning are generally used as multilamellar 3D structures ( Chen et al., 2011 ).
An alternative approach to producing 3D structures with electrospinning has been to add leachable
porogens to the system either in the form of co-electrospun fibers ( Baker et al., 2008 ) or salt particles
( Nam et al., 2007 ) to increase cellular infiltration and attachment throughout the inner regions of the
scaffold. Electrospun nanofiber scaffolds have been used in the regeneration of craniofacial bone de-
fects. Nonwoven electrospun PLLA nanofibers (mean diameter 775 ± 294 nm) ( Schofer et al., 2009 )
were collected in molds (diameter: 5 mm, height: 1 mm) and were implanted within same size defects
created in the dorsal part of the cranium ( Schofer et al., 2011 ). At 12 weeks post-surgery, PLA nanofiber
scaffolds and, more robustly, those applied together with 175 ng rhBMP-2 helped bridge the critical-size
cranial bone defects ( Figure s 10 . 3 G-I ) . In another study, nHAp/chitosan composite was electrospun to
prepare fibrous mats with mean fiber diameter in between 200-300 nm ( Liu et al., 2013 ). Three layers
of fiber mats were then held together and implanted within 5 mm × 5 mm rectangular full thickness
defects in the rat cranium. Better bone ingrowth and defect bridging was observed in the nHAp/chitosan
nanofibers as compared to chitosan nanofibers alone by the end of 20 weeks postimplantation.
Thermally induced phase separation together with porogen leaching has been used as another meth-
od to produce macroporous scaffolds with nanofiber features. In this technique, a polymer such as PLA
was dissolved in an organic solvent such as tetrahydrofuran and the resulting solution was separated
into distinct phases (PLA nanofibers and solvent) at low temperatures ( Gupte and Ma, 2012 ). The
solvent was then removed and the resulting nanofibrous scaffold (fibers on the order of 100 nm) was
collected. Interconnected porosity within the 3D polymer matrix was obtained by using sacrificial po-
rogen materials such as sugar or paraffin. Various studies have shown that this technique can produce
nanofibrous 3D scaffolds having interconnected micropores suitable for bone tissue engineering with a
variety of cell types and composite materials ( Li et al., 2002 ; Liu et al., 2009 ; Wei and Ma, 2006 ). This
technique was also used to produce anatomically shaped nanofibrous mandibular scaffolds. This was
achieved by forming a wax mold from the CT images using rapid prototyping technology and then by
using this mold to prepare the phase-separated nanofibrous PLLA scaffold as described above ( Chen
et al., 2006 ) ( Figure s 10.3 J-L).
10.4.3 NANOCARRIERS IN BIOACTIVE AGENT DELIVERY
Growth factors have important regulatory functions in bone regeneration, from induction of cell recruit-
ment and vascularization toward the defect area to differentiation of osteogenic precursors to mature
osteoblasts. Multiple growth factors act in time- and concentration-dependent manners during these
processes. Therefore, mimicking the growth factor dose and availability during bone regeneration is
central to achieving functional constructs.
Controlled release technology has developed substantially over the last decades and enables the en-
capsulation of growth factors in carrier structures to protect their bioactivity and to prolong and localize