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International Journal of Bioprinting 3D scaffold prevents tendon ossification

environments can induce structural reorganization into while concurrently evaluating systemic inflammatory and
ordered β-sheet configurations. This conformational immune responses.
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transition not only enhances SF’s mechanical properties but
also expands its biomedical applicability. Hydroxypropyl 2. Materials and methods
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cellulose (HPC), another biomaterial characterized by an 2.1. Formulation of bioink
abundance of hydroxyl groups, has been widely employed A total of 1.0 g SF, 1.0 g HPC, and TSPCs at a density of
to optimize SF performance. The incorporation of HPC 5×10 cells/mL were added to 10 mL phosphate-buffered
6
facilitates SF’s structural transition from random coils to saline (PBS). The mixture was gently homogenized using
β-sheets, thereby improving mechanical robustness and a magnetic stir bar at 100 rpm for 10 min to avoid cellular
structural stability. Notably, HPC-mediated molecular damage. To identify the optimal bioink composition, cross-
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interactions induce the formation of dual-network combinations of SF (0.5 g, 1.0 g, 1.5 g) and HPC (1.5 g, 1.0
hydrogels through this conformational shift, resulting g, 0.5 g) were systematically tested. The 1:1 SF–HPC ratio
in superior mechanical strength and elasticity. These (1.0 g SF + 1.0 g HPC) demonstrated superior rheological
36
SF–HPC composite hydrogels demonstrate significant properties and was selected for subsequent experiments.
potential for biomedical applications, particularly in For the SF control bioink, 2.0 g SF and TSPCs (5×10
6
leveraging their synergistic biocompatibility and controlled cells/mL) were dissolved in 10 mL PBS under identical
degradation profiles.
mixing conditions. During the experimental process, we
TSPCs, first identified in 2007, are multi-potent cells observed that the viscosity of the SF–HPC bioink was
37
capable of differentiating into tenocytes, chondrocytes, proportional to the concentration of HPC. However, when
and osteoblasts, playing a pivotal role in tendon repair. bioink with an SF–HPC ratio of 1.5:0.5 was used for 3D
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TSPCs address this challenge by undergoing tenogenic printing, the viscosity was too low to prevent scaffold
differentiation to promote collagen synthesis and collapse. Conversely, bioink with an SF–HPC ratio of
extracellular matrix deposition, thereby facilitating 0.5:1.5 tended to cause nozzle clogging during 3D printing.
functional tissue regeneration. Furthermore, TSPCs Therefore, a 1:1 SF–HPC ratio was determined to be the
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enhance tendon healing through immunomodulatory optimal formulation. TSPCs (No. CP-R165) used in the
effects, stimulation of tenocyte proliferation, and experiments were purchased from Procell Life Science &
acceleration of collagen remodeling. In tissue engineering, Technology Co., Ltd (China). Experiments were conducted
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TSPCs are frequently integrated with biomaterial scaffolds using cells at passages 2–3, as this stage is considered
to orchestrate neo-tendon formation. The combination of optimal for maintaining cellular functionality.
TSPCs with biocompatible scaffolds provides a biomimetic
microenvironment that supports cell adhesion, migration, 2.2. Rheological characterization of bioink
proliferation, and lineage-specific differentiation. For The rheological properties of the bioinks were analyzed
instance, decellularized tendon scaffolds functionalized using a modular advanced rheometer system (Thermo,
with collagen have demonstrated efficacy in promoting USA). Dynamic viscosity was measured under shear
−1
TSPC proliferation and tenogenic commitment, rate sweeps from 0.1 to 100 s at 37°C. Frequency-
ultimately enhancing the functional regeneration of dependent storage modulus (G’) and loss modulus (G’’)
damaged tendons. This synergistic approach exhibits were determined through dynamic frequency sweep tests
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substantial potential for advancing tendon tissue conducted over 30 min at 37°C.
engineering applications.
2.3. Three-dimensional bioprinting of tissue-
In summary, this study pioneers the development of engineered Achilles tendon scaffolds
a novel SF–HPC–TSPC bioink, formulated with SF as The SF–HPC–TSPCs bioink formulated in Section 2.1 was
the matrix, HPC as a reinforcing agent, and TSPCs as the employed to fabricate tissue-engineered Achilles tendon
cellular component. We first characterized the rheological scaffolds using a multi-nozzle 3D bioprinter (Discovery,
properties of this bioink and subsequently employed it Switzerland). A computer-aided design model of the
for 3D bioprinting of tissue-engineered Achilles tendon scaffold structure was first created, followed by loading
scaffolds. The printed scaffolds were rigorously evaluated for the bioink into the pressure-assisted nozzle cartridge of
mechanical properties, degradation behavior, and porous the bioprinter. Printing parameters were set to a nozzle
microstructure. In vitro investigations systematically pressure of 0.25 mPa and a deposition speed of 1.0 mm/s.
assessed the scaffold’s ability to regulate TSPC survival, Scaffolds with final dimensions of 2 × 3 × 15 mm (width ×
migration, proliferation, and tenogenic differentiation. height × length) were bioprinted onto sterile petri dishes
Furthermore, in vivo studies validated the scaffold’s under controlled environmental conditions (25°C; 60%
efficacy in preventing HO following Achilles tendon injury relative humidity).

Volume 11 Issue 4 (2025) 299 doi: 10.36922/IJB025210203