Page 286 - IJB-10-6

International Journal of Bioprinting Skin bioprinting: Keratinocytes and stem cells

cells and enhanced in vitro wound healing. These variability and inconsistency, making precise mechanical
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findings are consistent with the results of our 3D model. characterization challenging. According to Joodaki and
The bioprinted constructs containing HaCaT and ADSCs Panzer, the skin exhibits Young’s modulus of 1.1–210 kPa,
in co-culture revealed significantly higher cell viability on depending on the skin site, as determined in indentation
days 14 and 7 compared to day 1. In contrast, Alg/HA/ studies. The literature Young’s modulus of the upper
Gel constructs containing only HaCaT did not exhibit a skin layer is 0.11 kPa, whereas Young’s modulus of the
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statistically significant increase in cell viability from days 1 reticular dermis is much higher at 160 kPa. Furthermore,
to 14. Additionally, the live/dead assay indicated increasing the dermal skin layer provides additional stability and
live cell counts in constructs containing both HaCaT and significantly contributes to the stiffness of the skin. Since
ADSCs, whereas groups containing only HaCaT displayed our 3D model lacks a dermal layer of fibroblasts, which
increased live cell count only until day 7. Our study revealed would enhance the skin stiffness, our results are not
a mean survival rate of 88% for ADSCs within Alg/HA/Gel comparable to those of natural skin but fall within the
constructs and 92% within GelMA constructs throughout broad range aforementioned.
the experimental period, consistent with the findings of During the process of re-epithelialization, keratinocytes
Schmid et al., whereby Alg/HA/Gel bioinks resulted in proliferate and migrate towards the center of the lesion
good printability and ADSC survival, proliferation, and until the wound surface is covered by a monolayer of
differentiation. Moreover, Li et al. also demonstrated keratinocytes. To evaluate the re-epithelialization of the
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1,42
high ADSC proliferation and growth factor secretion after wound bed, the degree of cell migration out of the bioprinted
ADSC conjugation onto a GelMA hydrogel. 37 constructs to the well bottom was analyzed. For experimental
Mechanical stress from passing through the printing groups containing HaCaT and ADSC in co-culture, the
nozzle can cause cell damage or death. The shear- metabolic activity and viability of the cells at the well bottom
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thinning behavior of bioinks reduces viscosity during were significantly increased during the experimental period,
printing but maintains high viscosity when at rest. 26,39 Thus, independent of the selection of hydrogels. After 14 days,
the hydrogel could be printed at much lower pressure, 55% of the well bottom was covered by HaCaTs, which were
reducing the hydrodynamic stress on cells and improving initially printed in co-culture into Alg/HA/Gel constructs.
HaCaTs from co-cultured GelMA constructs covered 45%
cell viability. The high resting viscosity of hydrogels is of the well bottom. In a wound situation, it can be assumed
essential for the printability, shape-fidelity, and mechanical that keratinocytes can migrate from the bioprinted skin
stability of bioprinted constructs. The overall survival substitute onto the wound bed, potentially enhancing re-
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rate of all cells during the entire test period was at least epithelialization. Several studies have reported the stimulatory
75% with peak survival on day 7 at 82.5%, suggesting that effect of ADSC on the proliferation and migration of HaCaT
cells regenerated from the printing process and faced less cells. HaCaT proliferation and migration, under the influence
competitive pressure due to lower density in the bioprinted of ADSCs, were examined in vitro, using vitality assays and
constructs. To reduce mechanical stress and cell damage scratch tests. 34,36,43 Likewise, an in vivo mouse wound healing
or death, conical nozzles were used for bioprinting instead model validated the positive effects of ADSCs on HaCaT cells
of cylindrical ones, as the latter requires higher pressure and wound healing. 43
for printing at the same flow rate. Likewise, conical nozzles
can induce higher shear stress on the cells at a greater area In addition to the migrated HaCaT, a significantly
compared to cylindrical nozzles. 40 smaller amount of ADSCs migrated to the well bottom.
Seo et al. indicated that these migrated ADSCs could
The stiffness analysis confirmed the viscoelastic potentially differentiate into keratinocytes and further
properties of the bioprinted constructs, whereby the enhance re-epithelialization. 33
hydrogels primarily exhibit elastic properties. The
stiffness of the constructs was determined by the hydrogel The degradation assay displayed a significant decrease
composition, whereas cell type did not play a significant in the size of the biofabricated constructs throughout the
role. Biofabricated Alg/HA/Gel constructs reported a experimental period. In addition, cell-laden constructs
significantly higher elastic modulus compared to GelMA were significantly smaller on day 14 than the comparative
constructs, indicating that GelMA is softer than Alg/ cell-free group.
HA/Gel. However, the mechanical parameters of natural The degradation of Alg/HA/Gel and GelMA has been
skin vary widely and are influenced by patient-related reported previously. 26,27 Consistent with these findings, our
parameters, such as weight loss, body mass index (BMI), study revealed a higher degradation rate for Alg/HA/Gel
age, and the specific body site on which the skin was constructs compared to GelMA constructs, particularly for
obtained. These parameters introduce a high degree of cell-laden constructs. A degradation of 33% was observed

Volume 10 Issue 6 (2024) 278 doi: 10.36922/ijb.3925