Page 405 - IJB-10-3

International Journal of Bioprinting In situ thermal monitoring in bioprinting

(CELLINK, Gothenburg, Sweden), that is composed of for the selected bioink, pressure and print speed were
non-animal-derived polysaccharide components, alginate, set at 11 kPa and 20 mm/s respectively. For the second
and highly hydrated cellulose nanofibrils. It is a transparent campaign, due to the custom-made nature of the bioink,
bioink selected for its consistent and temperature- after several calibration sessions, for optimal printing, the
independent shear-thinning properties and its similarity to pressure and speed parameters were set at 7 kPa and 14
the extracellular matrix, both morphological and biological. mm/s, respectively.
In the second experimental campaign, bioprinting The printbed and printhead temperatures were fixed
was carried out by combining the neonatal human dermal at 20°C and 30°C, respectively, for all experimentations.
fibroblasts (nHDF; PromoCell, Heidelberg, Germany) For the second campaign, the printbed temperature acted
with a custom-made alginate-gelatin hydrogel (8% w/v for also as a thermal crosslinker for gelatin in the alginate–
both components), to test the capability of the approach gelatin hydrogels. Printed constructs were then ionically
of expanding its usefulness on different biomaterial and crosslinked at the end of the printing process by dropping
process conditions. The alginate-gelatin hydrogel represents on them a solution of calcium chloride (CaCl ).
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a water-rich network of hydrophilic polymers that absorb
water while maintaining their physical structure. This 2.4. 3D models
bioink reaches a liquid state at a temperature of 30°C and Square lattice patterns (20% infill density), commonly
due to the presence of gelatin shows a shear-thinning used in the EBB, were chosen as printing samples. At first,
behavior and undergoes gelation below room temperature, models of 10 × 10 × 1.6 mm were chosen (Figure 1). The 3D
forming a gel starting from a liquid state. models in STL format were created on SolidWorks software
(Dassault Systèmes SE, Vélizy-Villacoublay, France).
2.2. Cell culture In the first campaign, the same square lattice pattern
nHDFs were used for bioprinting experiments. For was repeated on each layer. In the second experimental
expansion, cells were plated in T75 flasks at a seeding campaign, i.e., combining bioink with the nHDF, we used
density of 3 × 10 cells/cm in 15 mL of PromoCell a different test where a different geometry was used at each
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2
Fibroblasts Growth Medium and then incubated at 37°C layer to produce a scaffold of 14 × 14 × 1.6 mm (the “step”
(with 5% CO ). For bioink preparation, the volume of model), just to show the impact of a temperature-based
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cell suspension needed for the desired cell count was reconstruction of the geometry, which allows in principle
centrifuged at 220 rpm for 3 min at 37°C. The resulting to observe/monitor the geometry of the last layer only.
pellet was resuspended and manually mixed in a 1:9 ratio
with the tested hydrogel. The bioink was prepared with a In order to produce this second model of interest, the
cell concentration of 1 × 10 cells/mL. G-code was modified using NC Viewer, an online open-
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source G-code programming application to obtain a model
Given the interest of our work in the technological where only a partial portion of the layer was printed at
innovation represented by the use of a thermal imaging layers 2, 3, and 4, as shown in Figure 2.
camera for in situ monitoring, no viability tests or follow-
up activities of the bioprinted constructs were carried
out. The cellular component was introduced inside the
hydrogels in order to monitor the extrusion process of a
bioink whose printing properties were representative of the
process under investigation, and thus with the rheological
properties of the hydrogels considered modified by the
presence of the cell suspension.

2.3. Bioprinting set-up
The process monitoring method was applied to a
pneumatic extrusion-based bioprinter, the BIO X
(CELLINK, Gothenburg, Sweden), that was used to
fabricate biocompatible scaffolds suitable for cells.
For the current work, general-purpose sterile high-
precision conical bioprinting nozzles with a nozzle internal
diameter of 0.41 (22 G) mm and a 32 mm conical nozzle
length were used. For the first campaign, according to the
manufacturer’s indications to obtain the best printability Figure 1. 3D representation of the standard model.

Volume 10 Issue 3 (2024) 397 doi: 10.36922/ijb.2021