Systematic multi-parameter optimization of an injectable thermosensitive CS/β-GP/HEC hydrogel for 3D cell culture...

Area of Science:

• Biomaterials Science
• Tissue Engineering
• Cell Biology

Background:

• Injectable thermosensitive hydrogels are crucial for minimally invasive 3D cell culture.
• Chitosan-based hydrogels offer tunable properties for biomedical applications.

Purpose of the Study:

• To systematically screen and optimize a chitosan (CS)/β-glycerophosphate (β-GP)/hydroxyethyl cellulose (HEC) ternary hydrogel system.
• To evaluate the suitability of optimized hydrogels as scaffolds for in vitro 3D cell culture.

Main Methods:

• A 2×2×2 factorial design was employed for systematic formulation screening.
• Gelation kinetics, injectability, and structural stability were assessed using a weighted scoring model.
• Biological evaluations included cell viability and proliferation assays in 3D hydrogel cultures.

Main Results:

• Two superior hydrogel formulations (G1 and G3) were identified, balancing rapid gelation at 37 °C with room-temperature injectability.
• The 3D hydrogels significantly enhanced cellular viability and spatial proliferation compared to 2D cultures up to Day 10.
• Huh7 cells formed dense tumor spheroids, and BMSCs established spatial networks within the hydrogel matrix.

Conclusions:

• A structured formulation-screening strategy was successfully implemented for developing injectable hydrogels.
• The CS/β-GP/HEC hydrogel system provides a stable, biomimetic matrix suitable for advanced in vitro 3D cell culture.
• These findings highlight the potential of optimized hydrogels for applications in regenerative medicine and drug screening.