CARTILAGE 3D PRINTING - 3D Bioprinting and Nanotechnology in Tissue Engineering and Regenerative Medicine - page 265

Cui et al. ( Cui et al., 2012b ) study, a cylindrical cartilage defect size, 4 mm in diameter and 2 mm
deep (extending to the subchondral bone), was filled in less than 2 min. Following simultaneous po-
lymerization with UV exposure during printing of each layer (approx. 18 m m each) the cell viability
was high ( ∼ 90%). In contrast, if the defect was first completely filled with PEGDMA, crosslinking,
took a minimum of 11 min, which also significantly reduced cell viability ( ∼ 63%) ( Kim et al., 2003 ).
Cross-linking during ink-jet printing also prevented cell settling at the interface between layers due
to gravity, as observed in other studies ( Kim et al., 2003; Sharma et al., 2007 ). Feasibility of control-
ling the 3D location of each printed cell can be very useful in reproducing the native organization of
cells and ECM. Examples of potential applications are: placing specific cell types (chondroprogeni-
tors and osteoprogenitors) in appropriate locations; placing zonally derived chondrocytes in their
proper zone ( Schuurman et al., 2013b ); and encapsulating cells in spatially varying ECM for better
defined zonal differentiation states ( Grogan et al., 2014; Grogan et al., 2013a; Klein et al., 2009;
Schuurman et al., 2013b ). After 6 weeks of culture, Cui et al. ( Cui et al., 2012b ) observed that the
printed PEG gel and chondrocytes produced cartilaginous neotissue with decreased collagen type
I and increased collagen type II (gene expression level), substantial GAG deposition (Safranin O
stain), and evidence of histologic integration with host tissue. By 6 weeks the compressive modulus
of the printed tissue approached 400 kPa, within the range of that reported for articular cartilage
( Lai and Levenston, 2010 ). Such mechanical properties may be suitable for surviving in a loaded
environment.
12.1.6 MAJOR CHALLENGES AND PITFALLS
The scientific and technical challenges facing successful bioprinting can be broadly classified into cel-
lular, bioink, and printing technology.
12.1.6.1 Cell source
One has to address the scarcity of obtaining normal autologous human chondrocytes in sufficient quan-
tities. One approach is to use autologous adult stem cells or progenitors that can be expanded into suf-
ficient quantities before differentiation into matrix-producing chondrocytes. Another approach is to use
an allogeneic cell source. Advantages of the former are lack of allogeneic response with lower risk for
regulatory approval. Advantages of the latter are a well-defined and characterized source of cells with
potential for off-the-shelf treatment. In addition to maintaining cell viability that survives the printing
process, one has to ensure an appropriate and enduring phenotype.
12.1.6.2 Scaffolds
Many challenges have to be overcome before printable scaffolds can be clinically useful. For successful
tissue regeneration, the scaffold must be biocompatible with minimal biological side effects upon deg-
radation or release of degradation products. The bioink has to rapidly undergo liquid-to-solid transition
to support efficient printing. Traditional methods used in commercial 3D printers, such as extrusion of
molten polymer or powder-based printing, are not likely to be compatible for obvious reasons. Novel
approaches to chemical or photocrosslinking biomaterials are presently being explored. Other chal-
lenges which the appropriate bioink must overcome include the properties to survive implantation and
postoperative use (such as mechanical strength), and the appropriate biochemical and biophysical cues
to maintain the phenotype of the printed cells.