RESEARCH SUMMARY
This study developed and benchmarked a multifunctional, thermoresponsive, electrically conductive hydrogel bioink intended for extrusion-based 3D bioprinting of electroactive tissue constructs and bioelectronic interfaces. The authors formulated a 12-member library of agarose/gelatin/hydroxypropyl cellulose (HPC) hydrogels with PEDOT:PSS as the conductive component (0.01–0.5% w/w), varying agarose:gelatin ratios while holding HPC constant (2% w/w). Formulations were screened for swelling and degradation behavior (PBS, 37 °C), rheology relevant to printing (three-interval thixotropy and oscillatory viscoelastic characterization), and quantitative print fidelity using line, grid, and multilayer cylinder geometries. Electrical performance was quantified by four-point probe conductivity and by electrochemical impedance spectroscopy (EIS) after printing the bioink onto screen-printed carbon electrodes. Biocompatibility was evaluated using A549 cells under multiple contact/encapsulation configurations (“on”, “under”, “in-between”, and “in”), with MTS, LDH, and live-cell fluorescence imaging over 24–72 h. A formulation containing 2% agarose, 4% gelatin, 2% HPC, and 0.1% PEDOT:PSS (“B2”) provided the most balanced performance: robust shear-thinning and recovery, high print consistency with stable multilayer stacking, physiologically relevant conductivity (~0.576 S/m), and no significant loss of cell viability relative to controls across embedding conditions. Overall, the work provides a systematic formulation strategy for conductive, thermoresponsive bioinks that can be printed and used immediately without post-printing functionalization, supporting applications in electroactive tissue engineering, biosensing, and wearable/soft bioelectronics.
Unconfined compression mechanical testing was performed using a CellScale UniVert mechanical tester equipped with a 100 N load cell to quantify how hydration state and physiological testing conditions affected the stiffness, strength, and energy absorption of the candidate bioink. Cylindrical hydrogel specimens were prepared by excising discs with a tissue hole punch (approximately 10 mm diameter and ~10 mm thickness). Samples were compressed at 15% strain per minute to a maximum of 30% strain with a 0.01 N preload while force and displacement were continuously recorded. Three conditions were compared: (i) dry/as-cast samples tested at ambient laboratory conditions, (ii) ‘wet’ samples hydrated overnight in PBS at 4 °C, lightly blotted, and equilibrated to room temperature prior to testing, and (iii) ‘submerged’ samples tested in a UniVert water-bath attachment filled with PBS and maintained at 37 °C to simulate physiological conditions. Stress–strain curves generated from UniVert data were used to calculate elastic modulus, ultimate compressive strength, and toughness (area under the curve). For the optimized B2 formulation, hydration increased mechanical performance: elastic modulus rose from 87.11 ± 19.59 kPa (dry) to 113.29 ± 10.03 kPa (wet) and 119.28 ± 9.80 kPa (submerged), while ultimate compressive strength increased from 12.68 ± 1.75 kPa (dry) to 15.16 ± 1.08 kPa (wet) and 17.09 ± 0.81 kPa (submerged). These UniVert measurements provided the study’s primary quantitative validation that the conductive, thermoresponsive bioink maintained and even improved compressive mechanical properties under hydrated and 37 °C testing—supporting its readiness for physiologically relevant 3D printed electroactive constructs.
