B14 - Biofabrication of 3D Printed Artery-on-chip with ECM based bioink

Introduction: : Organ-on-chip systems have emerged as transformative tools for modeling human diseases in vitro. Recapitulating the complex pathophysiology of vascular disorders like vasculitis remains an unmet challenge. Current models of vascular diseases often rely on oversimplified 2D cultures, which lack the multi-cellular architecture and human-specific microenvironment required to recapitulate disease progression and therapeutic responses. To address this gap, we present a 3D-bioprinted, medium-sized artery-on-chip platform that integrates primary human endothelial cells, smooth muscle cells, and fibroblasts into a physiologically structured construct. By optimizing the ECM based bioink formulation, tri-axial bioprinting, and post-printing processes, we aim to achieve spatially controlled cell spreading, phenotypic maintenance, and a confluent endothelial monolayer. Furthermore, we validate the model’s viability under long-term perfusion, which enables the study of inflammatory cascades and cytokine responses. This platform bridges the divide between oversimplified in vitro models and in vivo complexity, offering a robust tool for mechanistic studies and intervention development.

Materials and

Methods: : Primary vascular SMCs (2% alginate/3% gelatin/0.25 mg/ml Col IV) and fibroblasts (2% alginate/3% gelatin/0.5 mg/ml Col I) were triaxially bioprinted with a Pluronic F127 core (30% +100 mM CaCl₂). Arteries were extruded into 300 mM CaCl₂, followed by secondary crosslinking (20 mM CaCl₂ + 6 U/ml TG, 2 h) (Figure 1A). Alginate was leached (13.75 mM sodium citrate), and constructs were cannulated in 3D printed housing. Aortic ECs (20M/ml) were seeded in lumen on day 3. Cell morphology (IF), gene expression (qPCR), and barrier function (TRITC-dextran) were assessed. Artery viability under perfusion was evaluated by TUNEL for up to 21 days.

Results, Conclusions, and Discussions:: Z-stack and orthogonal imaging confirmed fibroblast and SMC spreading throughout the bioink following alginate leaching (Figure 1B). Temperature-controlled triaxial extrusion produced arteries with physiologically relevant wall thicknesses (200–400 μm). Collagen I/IV supplementation enhanced mechanical stability, enabling safe cannulation. Rapid crosslinking preserved layer-specific cell localization post-printing (Fig. 1C). Endothelial cells (20M/mL seeding density) formed a confluent monolayer by day 7. Ongoing work includes investigating endothelial barrier function through 20/70 kDa TRITC-dextran permeability assays (Figure 1D), evaluating long-term viability under physiological perfusion (0.5–20 dyn/cm²) in open and sealed chip configurations, and validating model fidelity via spatial single-cell transcriptomic comparison with native human arterial tissue.

Acknowledgements and/or References (Optional)::