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International Journal of Bioprinting Biocompatible materials and Multi Jet Fusion

Figure 8. Gene expression analysis. There are only two biological replicates in 28-day group, while the number of replicates of other groups is indicated in
Materials and methods. Note: ****P < 0.0001.
during culture in the construct; nonetheless, the change low viscosity provide a cell-friendly environment for longer
was statistically insignificant. culturing periods although their printability is usually
poor [47,48] . The amplitude sweep test proved that a lower
4. Discussion gelatin content corresponded with the occurrence of cross-

This work focused on the formulation of a bioink over at lower strain rates. Concomitantly, higher G’ values
composed of alginate, gelatin, and CCNC for meniscal 3D were observed for bioinks with a higher gelatin content,
bioprinting. Rheological analysis enabled us to determine which improves material strength but may result in poor
[49]
the optimal concentration of components. printability . Taking into account the entire viscosity
range and the temperature sweep test, the composition of
Firstly, the temperature sweep test was performed to bioink B with a gelatin content of 4.0% has the most suitable
establish an optimal gelatin content, since gelatin contributes rheological properties for 3D bioprinting and was chosen
the most to the temperature-dependent rheological for further analysis. More precisely, the pivotal impact on
properties of bioinks [37,39] . The onset of a significant decrease this selection includes the broad LVE region, the reasonably
in G’ was observed for all bioinks at 28°C, which is closely high G’ values, and the cross-over occurrence after a non-
related to the sol-gel transition temperature of gelatin [40,41] . rapid decrease of the storage modulus of bioink B.
Overall, gelatin is suitable for bioprinting at temperatures
below 28°C [41,42] . These results imply that the bioink should The amplitude sweep tests of bioinks with different
be cooled to at least 25°C before bioprinting. alginate concentrations (bioinks B, D, and E) revealed the
complexity of their viscoelastic properties and the inability
The shear-thinning behavior is another essential to predict their properties solely from the concentrations
property of bioinks, which allows for precise and stable of their constituents. The optimization of the alginate
prints [43,44] . Bioink viscosities in the range of 30 mPa·s concentration is not only crucial for the printability and
−1
to over 6·10 mPa·s are considered compatible for 3D mechanical properties of the construct, but also for cell
7
−1
extrusion bioprinting, and the viscosities of bioinks viability and proliferation [41,50] . Based on the rheological
A, B, and C were within this range [45,46] . An increased tests and the biological properties of alginate, the 0.75%
concentration of gelatin stiffens the bioinks within the alginate concentration was selected for further studies.
tested temperature range and ensures better printability and
stability of bioprinted constructs. However, an excessive Lastly, the rheological dissimilarities between bioinks
gelatin content may negatively affect the printing process with different CCNC concentrations (bioinks D, F, and G)
due to nozzle clogging or non-uniform bioink flow. Higher were assessed. This component has a significant impact
viscosity also causes cellular damage; hence, bioinks with on bioink reinforcement and the improvement of shear-

Volume 9 Issue 1 (2023) 8 https://doi.org/10.18063/ijb.v9i1.621