Page 471 - IJB-10-2

International Journal of Bioprinting Bioprinting with ASCs and bioactive glass

gelatin (Gel), and alginate+gelatin (Alg+Gel or AG) is composite AG gels made with higher glass percentages.
shown in Figure 3a, and the viscosity of B3 glass-modified Figure 4a shows the physical behavior of gels prepared
AG hydrogels with 1.25 wt.% (1.25G), 2.5 wt.% (2.5G), 5 with increasing glass content. It can be clearly seen that
wt.% (5G), and 10 wt.% (10G) glass content is shown in gel made with the most glass content behaves like a large
Figure 3b. The viscosity for all hydrogels was measured piece of crosslinked gel. For example, extruding or manual
at room temperature and before crosslinking with 0.1 M separation of 10G gel resulted in pockets of crosslinked
CaCl solution. At higher shear rates, modified hydrogels gel (pockets of gel chunks) being extruded or separated
2
with B3 glass slipped out of the plates (especially, 5G and from the hydrogel rather than a single continuous filament
10G) of the rheometer, and the test was stopped before the extrusion. This non-uniformity is believed to arise during
programmed shear rate of 100 s . Overall, all AG hydrogels the gel preparation process as pockets of alginate could
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irrespective of B3 glass addition showed a shear-thinning be crosslinked in a localized manner even before the
behavior with decreased viscosity at increasing shear rates. complete uniform mixing of hydrogel. As a result, during
As B3 glass is a fast-dissolving bioactive glass, it quickly the oscillatory sweep tests, chunks of gels were noticed to
2+
2+
starts to release Ca and Mg ions, which are divalent, break apart and come out of the plates at higher strain.
3+
+
and other ions including K , PO , Na , and B to the In addition to shear-thinning and solid-like behavior
3-
+
4
surrounding aqueous media. As the divalent ions initiate of hydrogels, the viscosity recovery of the hydrogel after
the crosslinking with alginate in the hydrogel, the viscosity removal of shear force is crucial in extrusion-based 3D
of hydrogels with B3 glass was significantly increased by bioprinting applications. As hydrogel is extruded through
several orders of magnitude (Figure 3b). The addition of 2.5 the nozzle, it suffers from higher shear stress and flows
wt.% B3 glass resulted in a significant and sharp increase through the tip because of a shear-thinning behavior. After
in hydrogel viscosity from 1.25 wt.% B3 glass addition deposition of a filament, the hydrogel should recover its
(from ~350 Pa·s to ~7000 Pa·s at 100 s ). The differences in molecular structure and viscosity to avoid spreading on the
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viscosity at t = 0 s observed for 2.5G, 5G, and 10G hydrogels substrate and withstand the weight of successive filaments
(~7000 Pa·s, ~10,000 Pa·s, and ~11,000 Pa·s at 100 s , that would be deposited on top of the current filament.
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respectively) were not as significant as viscosity differences To determine the recovery behavior of B3 glass-modified
between 1.25G and 2.5G hydrogels. These findings suggest AG hydrogels in a rheometer, hydrogels were initially
the likelihood of an optimal B3 glass concentration, ranging maintained under a steady-state shear rate of 0.1 s to
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between 1.25 and 2.5 wt.%, for AG hydrogels composed of 3 obtain a stabilized viscosity value. After reaching a steady-
w/v % alginate and 3 w/v % gelatin. state viscosity, the shear rate was increased to higher shear
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Amplitude sweep oscillatory tests were conducted rates (10 s and 100 s ) in two separate tests for a specific
on all samples to determine the viscoelastic behavior amount of time (30 s and 10 s, respectively), immediately
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of AG hydrogels modified with B3 glass. Figure 4 shows followed by a cooling period of 0.1 s shear rate. Viscosity
the variation of G’ and G’’ with strain percentage and of hydrogels recorded at different shear rates with time is
the linear viscoelastic region for each hydrogel. At low shown in Figure 5. The recovery data of two tests (at 10 s -1
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concentrations of glass (1.25%), there is no significant and at 100 s shear rates) were combined, and the time scale
difference in the storage modulus (G’) in comparison to was adjusted for a simplified representation. A significant
AG hydrogel (Figure 4b), whereas with the increase in drop in viscosity for all hydrogels at increased shear rates
glass concentration to 2.5% made a significant difference. can be clearly observed in Figure 5. Recovery time is defined
Figure 4c shows the variation in loss modulus (G’’) for all as the time taken for a hydrogel to attain its original steady-
gels investigated in this study. The transition from G’’>G’ state viscosity value from the reduced values at higher
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to G’>G’’ was more prominent in 2.5G, 5G (shown in shear rates. The shear rate of 100 s was used to mimic
Figure 4d and e), and 10G hydrogels clearly showing a the behavior of the hydrogel passing through the nozzle
viscoelastic solid-like behavior with a consistent behavior tip during extrusion. The results indicated a “near-zero”
of G’>G’’, indicating the increased stiffness of the modified recovery time for 2.5G and 5G hydrogels after application
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AG hydrogels with B3 glass. This is observed with a clearly of 100 s shear rate, which was evident from their step-
defined yield point (cross-over point of G’ and G’’). The function-like recovery to attain a constant viscosity at
shear stress values around the yield point for gels were 160 s, as shown in Figure 5. 1.25G and AG hydrogels
recorded as ~3400 Pa for 2.5G, ~3000 Pa for 5G, and ~1300 required 60 s and 90 s, respectively, to recover and attain a
Pa for 10G. It is also observed that the yield point occurs constant viscosity, as can be observed from the curvature
at lower strain for gels made with higher glass percentage. indicated by arrows in Figure 5. It was also observed that
This seems to be consistent with the lower shear stress hydrogels never truly recovered to 100% of their original
values. This could be because of the non-uniformity of the steady-state viscosity values (at t = 0 s) after the removal

Volume 10 Issue 2 (2024) 463 doi. 10.36922/ijb.2057