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Zhou, et al.
out from the needle, viscosity bounced back to 180 Pa∙s of the fraction of P4DA, implying that the excellent
within 5 s, which was 90% recovery of its initial viscosity. biodegradation properties of thermogels were not affected
The above results suggested that the hydrogels by the introduction of the covalent network.
possessed excellent shear-thinning and rapid recovery
properties, which were mainly attributed to the reversible 3.5. Printing of the dual-sensitive hydrogel inks
nature of physical bonding. Another worry on the thermo- To evaluate the printability of the dual-sensitive hydrogel
sensitive inks was that the solid hydrogels would transform inks, lattice structure was printed with DA00, DA20,
back into solutions at low environmental temperatures. DA40, DA60, DA80, and DA100. As shown in Figure 4A,
But fortunately, the thermogels exhibited a slow de-gelling multi-layer grids printed with DA00 and DA20 collapsed,
process; the viscosity of the system remained unchanged square holes in the structure transformed into circular ones
in 5 min even after the temperature was dropped from due to diffusion of filaments, showing their poor shape
37 to 25°C (Figure 2A). The self-supporting filament in retention. In contrast, constructs with the well-defined
the filament collapse experiments also demonstrated structure were obtained in experiments of DA40, DA60,
[50]
that the inks could hold the printed constructs before the DA80, and DA100. And their diffusion rate, a parameter
photo-crosslinking was constructed (Figure 4G). to evaluate ink’s shape fidelity was around 35%, much
[48]
3.4. Properties of the dual-sensitive hydrogel lower than that of DA00 (97.20%) and DA20 (78.80%).
In addition, diameters of filament (line width) were used
Successful printing is dependent not only on the to quantitatively assess the printability of different inks.
extrudability of inks but also on the ability to support DA00 and DA20 showed an unacceptable line width
the printed constructs. However, the thermo-sensitive (1.37 and 0.93 mm), which was three-fold larger than the
hydrogel was a physical hydrogel with weak mechanical needle diameters (0.31 mm). Nevertheless, the line width
strength. The maximum value of G’ in temperature sweep of DA40, DA60, DA80, and DA100 was, respectively,
was only 500 Pa (Figure 2C), indicating poor shape determined to be 0.44, 0.47, 0.46, and 0.53 mm, which
retention of the hydrogels. To compare the mechanical was quite close to the diameters of needle. There was no
strength of hydrogels after photo-crosslinking, G’ of doubt that the printability of the inks was improved by
all samples was evaluated using frequency sweep tests. the addition of P4DA, which was attributed to enhanced
In Figure 3D, G’ (at 1 rad/s) was increased from 90.7 mechanical strength due to the formation of covalent
Pa of DA00 to 7493.1 Pa of DA100, indicated that the networks. Therefore, inks with P4DA fraction above
mechanical strength was enhanced by the introduction of 40 wt.% were expected to be used in further experiments.
photo-crosslinking. Furthermore, this was also confirmed Next, DA40 was chosen as a model to explore the
by the results of tensile tests, DA100 reached a tensile influence of printing parameters on the inks’ printability.
modulus of 21.9 kPa (Figure S4) but DA00 was too weak The needles with different diameters were used to print the
to form a specific shape for tests. In addition, the storage inks. The results showed that the inks could be extruded
and tensile modulus tended to be higher when more P4DA from all needles with diameters from 0.16 mm to 0.53 mm
was formulated into the inks. Therefore, by varying the (Figure 4B). However, the smaller the diameters of the
fraction of P4DA in the inks, the hydrogels’ mechanical needle, the higher pressure was needed to extrude the inks.
strength can be tuned to satisfy a range of applications. Besides, all the results showed a line width larger than their
The swelling properties of the samples were needle diameters (Figure 4E), which was attributed to the
investigated by immersing them in deionized water. spreading of the weak thermo-sensitive hydrogel before
DA00 (848.2%) and DA20 (844.26%) had a high photo-crosslinking. Filament interval, representing the
swelling ratio because of the lack of effective covalent distance a strand needed to span between filaments from
network formation. However, the swelling ratio of DA40 the previous layer, is another important printing parament.
was decreased (771.7%), which was attributed to a As shown in the filament collapse study (Figure 4G), a
denser hydrogel mesh formed by chemical crosslinking. sagging filament was observed between a large gap, which
As the fraction of P4DA further increased, the swelling would result in poor shape fidelity. However, details of the
ratio of DA60, DA80, and DA100 was decreased to printed constructs would disappear when a small interval
559.7%, 451.3%, and 430.0%, respectively. The higher was chosen due to diffusion of filament into the deep part
swelling ratio of physical hydrogels was associated with of the construct (Figure 4C and F). For better printing
low structural stability . The reduced water absorption results, a needle with 0.31 mm diameter and a filament
[51]
of dual-sensitive hydrogels vouched for good shape interval of 1.2 mm was chosen for further experiments.
retention under cell culture over a long time (Figure 3G). With optimized parameters, a 10-layer lattice was printed
The enzymatic degradation properties of the hydrogels and displayed in Figure 4H. The printed grid exhibited
were then evaluated. As displayed in Figure 3E, all the high shape fidelity; moreover, it could be held by hands,
samples showed close degradation profiles regardless demonstrating the ink’s ability to print large structures.

International Journal of Bioprinting (2021)–Volume 7, Issue 3 147