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Nazia Mehrban, Gui Zhen Teoh and Martin Anthony Birchall

even more complexity to an already modified system. The main issue in using this approach is matching
the printable properties of the separate materials or
4.2 Natural Materials selecting a bioprinting technique which would allow

In contrast, natural materials, although inferior to both materials to be printed simultaneously under dif-
synthetic hydrogels in terms of controlling gelation ferent conditions. Although the latter adds another
kinetics and mechanical strength, are able to chemi- level of complexity to printing 3D biocompatible
cally and physically mimic native extracellular matrix scaffolds, it is a branch of bioprinting that is currently
(ECM). Collagen is the most abundant component of being explored [86] .
ECM [73] . It is widely used in tissue engineering appli- 5. Cell Encapsulation in Hydrogels for Printa-
cations and contains cell-guiding chemical cues, such
as the cell adhesion peptide sequence arginine-glycine- ble Bioinks
aspartic acid (RGD) [74] . However, although it is wide- The choice of cells for 3D bioprinting is often based
ly used as a bioprinting material, collagen is an un- on the type of tissue being created. However, as tis-
likely gold-standard candidate as it contracts and does sues and organs are composed of multiple cell types
not retain its original shape. which have a range of specific functions, it is likely
Hyaluronic acid (HA), is also a naturally derived that the bioprinting requirement will be for a mixture
material which does retain its shape and is already of cells. Current methods predominantly involve
used clinically [75] . HA forms very soft gels but can be printing individual cell types in specific patterns, de-
modified and crosslinked using a variety of methods signed to mimic native tissue cell distribution [87] . Al-
including the UV method described in Section 4.1 [76] though cells have been printed in single drops, with
and thiol-modified HA using gold nanoparticles [77] to each drop containing one or two cells [88] , it is currently
increase its stiffness. Similarly, fibrin is already used not possible to print individual cells reliably. This is
in surgery as a haemostatic agent and sealant [78,79] . The not an issue as long as large cell agglomerates (clus-
added complexity with fibrin is that it crosslinks ters of cells large enough to cause cell death at the
through the addition of thrombin. However, it can centre of the cell mass) can be avoided and cell-to-cell
produce mechanically stable hydrogels and has been contact can be maintained. The size of these agglo-
blended with other gels for bioprinting purposes [80] . merates will depend on the type of cells used and the
Some natural gels are difficult to print, not because ease with which nutrient and waste exchange can oc-
they form soft gels as described in the earlier exam- cur at the centre of the mass.
ples, but because their gelation properties are unde- For a more efficient system, resembling a native 3D
sirable. Gelatin is one such material. It forms a gel environment, a material-cell composite ink would be
easily by temperature control but has a melting tem- more suitable. The ability to encapsulate cells within
perature of 30–35°C [81] , which is below the standard the material as it is being printed allows researchers to
physiological temperature of 37°C. Similarly, alginate create a more tissue-like environment compared with
produces gels easily through cation crosslinking, but creating a 2D construct first onto which cells are then
unless it is modified with motifs that can guide cells to seeded [89] . With hydrogels this has been attempted
adhere, proliferate and differentiate, it is relatively with some success [90] , creating cell-laden constructs
inert [82] . that contain microvascular networks [91] and are able to
4.3 Hybrid Materials integrate well with native tissue [22] . Combining cells
with hydrogels is a delicate balance of maintaining
An alternative approach to producing scaffolds with high cell viability whilst ensuring that there are not
desirable properties is to create a hybrid. A study on too many cells in the gel to cause hyperplasia or
methacrylated hyaluronic acid combined with metha- apoptosis, either by optimising the number of cells
crylated gelatin showed that not only could cell viabil- added at the loading stage of the process or by con-
ity be maintained but by varying the concentrations of trolling the rate of cell proliferation post-printing [13] .
the two materials, the stiffness and viscosity of the When using hydrogels with cells, there are a num-
hybrid could be controlled [83] . Other researchers have ber of factors which could cause cell death. One of the
used a similar approach to bioprint scaffolds for a most obvious causes is the method selected for gela-
range of uses, including cartilage engineering [84] and tion. During crosslinking or temperature-based gela-
to tune material properties for a range of scaffolds [85] . tion the cell viability could be substantially affected [92] .

International Journal of Bioprinting (2016)–Volume 2, Issue 1 11