Mimicking Transmural Helical Cardiomyofibre Orientation Using Bouligand-like Pore Structures in Ice-Templated Collagen Scaffolds

Abstract

The helical arrangement of cardiac muscle fibres underpins the contractile properties of the heart chamber. Across the heart wall, the helical angle of the aligned fibres changes gradually across the range of 90-180°. It is essential to recreate this structural hierarchy in vitro for developing functional artificial tissue. Ice templating can achieve single-oriented pore alignment via unidirectional ice solidification with a flat base mould design. We hypothesise that the orientation of aligned pores can be controlled simply via base topography, and we propose a scalable base design to recapitulate the transmural fibre orientation. We have utilised finite element simulations for rapid testing of base designs, followed by experimental confirmation of the Bouligand-like orientation. X-ray microtomography of experimental samples showed a gradual shift of 106 ± 10°, with the flexibility to tailor pore size and spatial helical angle distribution for personalised medicine.

Keywords: anisotropic porosity; cardiac tissue engineering; freeze drying; polymer processing.

Figure 1

The base design in ice templating. (a) Overview of the cardiomyofibre orientation in native heart. (b–d) Base designs in ice-templating method to introduce 1D and 2D thermal gradients for freezing the slurry, where orange and blue represent different thermal conductivity. (e) Our Bouligand-like orientation design introduces 3D thermal gradient and (f) the experimental confirmation for the base design.

Figure 2

The wedge base design. (a) Overview of the wedge base design with illustration of $\beta$ as the slope angle. (b) The analysis plane for orientation analysis. (c) The snapshot of IFF positions at 500 s intervals for the 60° wedge base mould. (d) When $\beta$ increases from 15° to 60°, the streamlines on the analysis plane at the time of freezing completion. (e) The linear regression results of the IFF slope during the simulation when $\beta$ changed between 15° and 60°. (f,g) The temperature profile and phase separation diagram as the simulation progressed for the 60° wedge base mould.

Figure 3

The Bouligand-like orientation design. (a) The overview of the Bouligand-like orientation mould. (b) Details of the base topography. The base angle was defined as $\theta$, and pore orientation deviation from the vertical alignment was defined as $\phi$. (c,d) The schematic diagram of the temperature profile and IFF movement as the simulation progressed. (e) The region of interest for orientation analysis was highlighted in red. (f) Within the region of interest, the streamlines above a certain base and its corresponding pore orientation distribution. (g,h) The 3D distribution of streamlines within the prism region for (g) the equal width design (prototype mould) and (h) the unequal width design (refined mould).

Figure 4

The structural analysis of the ice-templated collagen scaffolds. (a,b) The reconstructed $\mu$CT images and corresponding pore orientation distribution of the samples fabricated using the prototype moulds. The area of interest was 1.2 mm $\times$ 1.2 mm at the centre of the prism region. (c,d) The reconstructed $\mu$CT images and corresponding pore orientation distribution of the samples fabricated using the refined moulds. The area of interest was 2 mm $\times$ 2 mm at the centre of the prism region.

Figure 5

Further characterisation of the refined mould samples. (a) Illustration of the volume of interest position. (b) The pore orientation distribution when moving away from the base. (c) The overview of maximum orientation shift within the scaffold using two freezing protocols (holding the temperature at −5 °C and −10 °C).