Page 480 - IJB-9-6

International Journal of Bioprinting Bioprinting cell-laden protein-based hydrogel

beyond this temperature, hydrogen bonds weaken, and wavelength and exposure times of 15, 38, and 60 s) and
the helices disappear, leading to a gel-sol transition. As a determined their ideal combination in the case of optimal
result of this well-defined feature, gelatin has proven to be stiffness for cartilage bioengineering. Human chondrocyte
a versatile additive for the formulation of a wide variety of collagen type II–alpha-I–Gaussia luciferase reporter
composite PBHs. Analogous to gelatin, collagen can also system (HuCol2gLuc) (cell density: 2 × 10 cells/mL) were
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form gels at different temperatures and pH levels [104,105] . embedded within the GelMA/HAMA bioink (15% w/v and
Compared with native tissues, its secondary helical 2% w/v), and the bioink was printed utilizing a pneumatic
conformation initiates a sol-gel transition for bioprinting extrusion printing device. It was revealed that the HAMA
but can commonly cause mechanical weakness. Hence, addition to GelMA enhanced chondrogenesis, compared
the molecular assembly of collagen is not controlled to GelMA (15% w/v) alone. Two ratios of GelMA/HAMA
sufficiently, and therefore, native structural hierarchy is bioinks (1:1 and 2:1) with different stiffness (32 and 57.9
not present [106] . Moreover, fibrin and silk fibroin hydrogels kPa) were assessed for cellular mobility and survival. High
display favorable outcomes and have unique advantages cellular viability (≥ 90%) was achieved for both bioprinted
in certain applications, including ease of preparation materials on all days. Furthermore, the GelMA/HAMA
and manipulation for fibrin and superior toughness and bioink with 1:1 ratio had considerably more fluorescent-
thermal stability for silk fibroin [107-110] . Secondly, PBHs labeled mobile cells (~2.5) compared to the 2:1 one (~1),
should have appropriate mechanical features, especially and the cells also moved further (~26 μm) and faster
elastic stiffness, which facilitate mechanical signaling (~1 µm/min) in the 1:1 ratio. All in all, the softer printed
to the cells for guiding differentiation, proliferation, construct (1:1) showed a higher level of cellular mobility
and ECM deposition [102] . In the case of stiffness, bioinks compared with the stiffer one (2:1). Notably, if multiple
with high stiffness or rigidity can inhibit the movement, kinds of cell are contained in the PBHs, the characteristics
growth, and differentiation of cells, whereas those with of the PBHs should possess tenability to accommodate the
high softness are able to provide sufficient support for various cellular requirements and allow spatiotemporal
the encapsulated cells [111-113] . Concerning these subjects, a control if required.
team of scientists [114] optimized the mechanical stiffness
of alginate/gelatin bioinks (crosslinking approach: ionic 3.1.2. Biodegradation
crosslinking with 2% w/v CaCl for 10 min) so as to Bioinks that degrade too rapidly cannot provide adequate
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improve ECM mineralization and cell organization for mechanical stability for long-term tissue development,
bone TE. With the aid of extrusion bioprinting, alginate/ while those that degrade too slowly may hinder tissue
gelatin bioinks (4.1% w/v for gelatin and 0.8% as well as growth and ECM remodeling. In addition to being an
1.8% w/v for alginate) loaded with P3 human mesenchymal essential feature for ECM deposition, PBHs’ optimal
stem cells (hMSCs) (cell density: 1.67 × 10 , 5 × 10 , and degradation is indeed necessary for cell proliferation and
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15 × 10 cells/mL) having various stiffness were printed. mobility in the gel as well as the nearby host tissue [116] . In
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They illustrated that soft scaffolds (0.8% w/v alginate, order to prolong the degradation rate of GelMA hydrogel,
stiffness: 0.66 ± 0.08 kPa) possessed higher content glycidyl-methacrylated HA (GMHA) was introduced
of deoxyribonucleic acid (DNA) at day 28, enhanced into this system to obtain a novel bioink with a suitable
expression of collagen type I alpha-II (6.7-fold increment degradation rate for cartilage regeneration applications.
from day 1 to day 28), increased alkaline phosphatase (ALP) Employing extrusion bioprinting, GelMA/GMHA bioinks
activity at day 28, and stimulated osteogenic differentiation (crosslinking approach: photo-crosslinking with 0.03% w/v
in comparison with the stiff ones (1.8% w/v alginate, 5.4 LAP) with various concentrations (7% w/v for GelMA and
± 1.2 kPa). Moreover, considerably less mineralized tissue 3% and 5% w/v for GMHA) encapsulating tonsil-derived
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was formed in stiff constructs than in soft ones at day 42 MSCs (cell density: 10 × 10 cells/mL) were printed, and
(22.6 ± 6.0 mm versus 43.5 ± 7.1 mm ) (Figure 3A). The the achieved constructs were subcutaneously implanted
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mineral formation rate in the soft bioprinted scaffold was into female Bagg albino (BALB)/c nude mice models so as
significantly higher than the stiff one at days 28–35 and to observe the hydrogels’ degradation characteristics after
35–42. Interestingly, cells in soft scaffolds displayed a 3D 3 weeks. It was illustrated that the 7% GelMA/5% GMHA-
cellular network in the mineralized matrix and osteoblast- printed scaffold was fairly stable within the physiological
and early osteocyte-related gene expressions at day 42. environment and preserved its shape well with a low
Within a pioneering investigation, Martyniak et al. [115] rate of degradation, allowing for tonsil-derived MSCs’
developed gelatin methacryloyl (GelMA)/hyaluronic acid chondrogenesis. Cells had high viability in the implanted
methacrylate (HAMA) bioinks (crosslinking approach: hydrogel, and cartilage-like tissues, that were regenerated
photo-crosslinking with 0.05% w/v lithium phenyl- over time, could be observed. Furthermore, enhanced
2,4,6-trimethylbenzoylphosphinate [LAP] at 405 nm expression of collagen type II and formed hyaline matrices

Volume 9 Issue 6 (2023) 472 https://doi.org/10.36922/ijb.1089