Oscillating perfusion of cell suspensions through three‐dimensional scaffolds enhances cell seeding efficiency and uniformity

Abstract

We developed a bioreactor for automated cell seeding of three-dimensional scaffolds by continuous perfusion of a cell suspension through the scaffold pores in oscillating directions. Using quantitative biochemical and image analysis techniques, we then evaluated the efficiency and uniformity of perfusion seeding of Polyactive foams as compared to conventional static and spinner flask methods. Finally, we assessed the efficacy of the perfusion seeding technique for different scaffolds and cell types. Perfusion seeding of chondrocytes into Polyactive foams resulted in "viable cell seeding efficiencies," defined as the percentages of initially loaded cells that were seeded and remained viable, that were significantly higher (75 +/- 6%) than those by static (57% +/- 5%) and spinner flask seeding (55% +/- 8%). In addition, as compared to static and spinner flask methods, cells seeded by perfusion were respectively 2.6-fold and 3.8-fold more uniformly distributed and formed more homogeneously sized cell clusters. Chondrocytes seeded by perfusion into Hyaff-11 nonwoven meshes were 26% and 63%, respectively, more uniformly distributed than following static and spinner flask seeding. Bone marrow stromal cells seeded by perfusion into ChronOS porous ceramics were homogeneously distributed throughout the scaffold volume, while following the static method, cells were found only near the top surface of the ceramic. In summary, we demonstrated that our cell seeding perfusion bioreactor generated constructs with remarkably uniform cell distributions at high efficiencies, and was effective for a variety of scaffolds and different mesenchymal cell types.

Influence of flow rate and scaffold pore size on cell behavior during mechanical stimulation in a flow perfusion bioreactor

“…Differences in geometrical configurations of perfusion bioreactor systems and the complex internal architecture of scaffolds (Bancroft et al, 2002;Bonvin et al, 2010;Wendt et al, 2003;Zhao and Ma, 2005) makes correlating observed biological outcomes on the basis of inlet fluid flow-rate very difficult. Calculation of the  within systems allows comparisons to be made on a single representative scale.…”

Cardiac tissue engineering using perfusion bioreactor systems

“…Recently, we adapted the methods established by Wendt et al 21 for cell seeding onto PGS by perfusion of a cell suspension through the scaffold voids. We determined the proper flow rate to use in order to achieve a dense, uniform spatial distribution of cells throughout the scaffolds.…”

Skeletal tissue engineering using embryonic stem cells

“…More homogeneous [200] or less uniform cell distribution [199] was also observed. Perfusion seeding increased the cell seeding efficiency and uniformity [199]. In our studies, static seeding on CM scaffolds was inefficient and not homogeneous.…”

“…Some studies claim more efficient seeding [198], whereas others found lower seeding efficiency in a spinner flask compared to static seeding [199,200]. More homogeneous [200] or less uniform cell distribution [199] was also observed. Perfusion seeding increased the cell seeding efficiency and uniformity [199].…”