Expansion of human amniotic fluid stem cells in 3-dimensional fibrous scaffolds in a stirred bioreactor

Human amniotic fluid stem cells (AFSCs) are emerging as an important cell source for tissue engineering and regenerative medicine due to their easy accessibility and broad multi-potentiality. In clinical applications, a large number of human AFSCs are required, which cannot be provided in conventional 2-dimensional (2-D) culture systems. To address this issue, the expansion of human AFSCs in 3-dimensional (3-D) polyethylene terephthalate (PET) scaffolds in a stirred bioreactor was evaluated. The results showed that 3-D PET scaffold with in vivo-like environment and a large specific surface area for cell adhesion promoted cell expansion (66-fold vs. 38-fold) compared to 2-D culture. A dynamic fibrous bed bioreactor (FBB) was used to expand AFSCs to reach a high cell density of 3.2 × 106 cells/mL. The bioreactor-expanded cells maintained clonogenic ability and high levels of expression (95.5-99.8%) of characteristic stem cell surface makers, including CD29, CD44, CD90 and CD105. The differentiation of bioreactor-expanded AFSCs into osteogenic and adipogeneic lineages was demonstrated with Alizarin red S and Oil Red O staining, respectively, and further confirmed by reverse transcriptase polymerase chain reaction (RT-PCR) analysis. This study demonstrated the feasibility of using the FBB to mass-produce human AFSCs for potential applications in tissue engineering and regenerative medicine.