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International Journal of Bioprinting Bioprinted cell-laden hydrogel for tracheal application

for homogeneous cartilage formation, difficulty in inflammation after in vivo implantation, caused by local
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customizing tracheal morphology, precise regeneration of inflammatory reactions resulting from surgical trauma
C-shaped tracheal cartilage, local inflammatory reactions and introduced materials, and further exacerbated
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caused by surgical trauma, introduced scaffolds that by respiratory microbiota. 19-21 The tracheal ciliated
inevitably compromise the structure and functionality of epithelial tissue layer plays a crucial role in protecting
TETC, and infection caused by respiratory microbiota cartilaginous tissue against inflammatory and bacterial
9
that may further lead to TETC deterioration. 10,11 Therefore, invasion. 22-24 Our previous study demonstrated that re-
there is still a need for new strategies to construct TETC epithelialization promotes TETC regeneration, but self-
with uniform chondrocyte distribution, customized migration is not a reliable and effective method, especially
cartilage shape, as well as anti-inflammatory and anti- for long-segmental tracheal defects. Existing studies have
3
bacterial functions. attempted to address this issue by preparing a sinusoidal-
The routine method for constructing a TETC involves patterned tubular mesh or developing a porous O-shaped
9,17
seeding chondrocytes onto a porous scaffold. However, scaffold ring with anti-inflammatory effects. However,
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uneven cell distribution and chondrocyte waste caused these studies neglected the importance of anti-bacterial
by the turbulent flow of the cell seeding suspension function. Therefore, the current study aimed to develop
limit its further application. 13-15 Three-dimensional (3D) a hydrogel that could be 3D-bioprinted with cells to
bioprinting is an emerging technology that provides generate a precise C-shaped tracheal ring and exert both
precise control over the fabricated constructs, including anti-inflammatory and anti-bacterial functions to enhance
cell distribution and structure. Park et al. presented TETC formation and tracheal restoration after orthotopic
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an advanced extrusion-based 3D-bioprinting strategy tracheal transplantation.
involving a two-step printing process: (i) printing a Icariin (ICA) is a prenylated flavonol glycoside
porous bellows framework and (ii) printing the cartilage extracted from Epimedium that has been shown to have
rings. In our previous study, we successfully developed favorable anti-inflammatory and chondroprotective effects
17
a 3D-bioprinted tracheal tissue incorporating O-shaped by modulating autophagy and apoptosis. 25,26 In addition, a
cartilage, providing further evidence of the capability study has suggested that ICA could be used as an alternative
of 3D-bioprinting technology to achieve uniform cell to growth factors for TETC regeneration due to its
distribution and precise shape control. However, recent chondrogenic effects. Chitosan (CS) is a natural-derived
18
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advancements from our research group have demonstrated polysaccharide approved by the U.S. Food and Drug
that a C-shaped cartilage configuration can more accurately Administration (FDA) for pharmaceutical applications. It
mimic the structure and function of native tracheal has excellent anti-bacterial ability attributed to the binding
cartilage, resulting in enhanced tracheal cartilage formation of its positively charged NH groups to negatively charged
3+
and restoration of tracheal defects. Notably, the mentioned bacterial surfaces. The anti-bacterial effect of CS is essential
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study employed a method involving the direct injection of in preventing infection due to the bacterial environment
cell-loaded hydrogel into customized C-shaped molds to of the tracheal lumen. However, poor reproducibility and
create the C-shaped cartilage. Nevertheless, this approach mechanical properties are insurmountable restrictions of
6
presents limitations when constructing long-segmental CS-based hydrogels used as bioinks. Gelatin methacryloyl
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tissue-engineered tracheal constructs with patient-specific (GelMA) hydrogel has gained increasing attention due to
shapes. Moreover, the mold-based method is intricate, time- its ability to crosslink and form hydrogels with tunable
consuming, and associated with a low success rate, thereby mechanical properties and excellent biocompatibility,
greatly restricting its practicality in applications. Therefore, mimicking the microenvironment of the native extracellular
it remains crucial to investigate the feasibility of utilizing matrix (ECM). In addition to acting as a carrier of cells,
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3D-bioprinting technology to construct tissue-engineered GelMA can also serve as a controlled drug delivery system
tracheal constructs with precise C-shaped cartilage for the administration of various active substances, such as
structures. Additionally, compared to 3D bioprinting a drug molecules. 16
trachea-mimetic cellular construct, the process of 3D In this study, we propose to enhance the anti-
bioprinting a single C-shaped cell-laden hydrogel ring inflammatory and anti-bacterial properties of GelMA by
requires less printing time, which can contribute to incorporating both ICA and CS. Our objective is to develop
improved chondrocyte viability and survival. 17
an ICA/CS/GelMA hydrogel that can be utilized in the
However, despite advancements in developing TETC fabrication of a TETC. We hypothesize that this hydrogel
using various techniques, including 3D bioprinting and can be pre-loaded with chondrocytes and subjected to a
scaffold-based approaches, several challenges remain. 3D-bioprinting process to create a TETC with a precise
One critical issue is the susceptibility of TETC to C-shaped ring. Doing so, we can address several challenges

Volume 10 Issue 1 (2024) 161 https://doi.org/10.36922/ijb.0146