Tissue engineering aims to regenerate or create damaged or lost organs and tissues by utilizing biodegradable scaffolds, cells and growth factors. One promising tissue engineering strategy involves seeding cells into a porous scaffold and culturing it in vitro, which is followed by implantation into the defect site. Cell seeding should result in a uniform distribution of cells inside the scaffold - otherwise the functionality and mechanical properties of the engineered construct can be compromised. Also a high seeding efficiency is appreciated to avoid wasting valuable cells and to enable faster tissue formation.

The aim of this study was to test six different cell seeding methods in order to find an optimal method for supercritical carbon dioxide (ScCO2) processed scaffolds. Of these scaffolds, one was a copolymeric poly(L-lactide-co-ε-caprolactone) 70/30 (PLCL) scaffold, whereas the other one was a composite of PLCL and β-tricalcium phosphate. The functionality of the cell seeding methods was verified with two scaffold types that were manufactured from poly-L/D-lactide 96L/4D (PLDLA 96/4) fibers. In addition, a novel cell seeding model utilizing iron-labeled microspheres with diameters of 15 µm and 100 µm was proposed. Adipose-derived stem cells (ASC) were used in the experiments due to their potential in hard-tissue engineering.

The study revealed that the microsphere seeding model is functional, offering useful information about the seedability of the ScCO2 processed scaffolds. The microsphere distributions were noticed to be more uniform compared to the corresponding cell seeding results. The microsphere model also suggested more challenging seedability of the composite scaffolds compared to the PLCL substrates. Applying micro-computed tomography (micro-CT) imaging and seeding ASCs fed with iron oxide nanoparticles, it was noted that the uniformity of the cell distribution in ScCO2 processed PLCL scaffolds can be enhanced by forcing the cell suspension into the scaffold with a syringe.

Due to technological limitations, evaluating cell seeding in the composite scaffolds was more challenging. However, the cell experiments supported the microsphere model, indicating a more difficult seedability compared to the PLCL scaffolds. A static pipetting of cells on top of the PLDLA fabrics was enough to provide desirable cell distribution and cell numbers in those scaffold types.