Evaluation of cell survival in different 3D-printed geometric shapes of human iPSC-derived engineered heart tissue

Abstract

Objectives

Engineered Heart Tissue (EHT) is a promising tool to repair heart muscle defects and can additionally be used for drug testing. Due to the absence of an in vitro vascularization, EHT geometry crucially impacts nutrient and oxygen supply by diffusion capacity. We analyzed cardiomyocyte survival in different EHT geometries.

Methods

Different geometries with varying surface-area-to-volume-ratios were calculated (structure A (Ring) AS/V = 58.47 mm²/440 μL³, structure B (Infinity) 25.86 mm²/440 μL³). EHTs were generated from hiPSC-derived cardiomyocytes (4 × 10⁶) and a fibrin/thrombin hydrogel. Cell viability was evaluated by RT-PCR, cytometric studies, and Bioluminescence imaging.

Results

Using 3D-printed casting molds, spontaneously beating EHTs can be generated in various geometric forms. At day 7, the RT-PCR analyses showed a significantly higher Troponin-T value in ring EHTs, compared to infinity EHTs. In cytometric studies, we evaluated 15% more Troponin-T positive cells in ring (73% ± 12%), compared to infinity EHTs (58% ± 11%, p = 0.04). BLI visualized significantly higher cell survival in ring EHTs (ROI = A: 1.14 × 10⁶ p/s and B: 8.47 × 10⁵ p/s, p < 0.001) compared to infinity EHTs during longitudinal cultivation process.

Conclusion

Use of 3D-printing allows the creation of EHTs in all desired geometric shapes. The geometry with an optimized surface-area-to-volume-ratio (ring EHT) demonstrated a significantly higher cell survival measured by RT-PCR, Bioluminescence imaging, and cytometric studies using FACS analysis.