Novel cell immobilization method utilizing centrifugal force to achieve high-density hepatocyte culture in porous scaffold

Abstract

Cell seeding is one of the key procedures in the construction of tissue-engineered organs. In our previous efforts to create a bioartificial liver, high-density cultures of hepatocytes (>1 x 10(7) cells/1 cm(3)-substrate) and long-term maintenance of metabolic function were achieved with a packed-bed reactor utilizing porous poly(vinyl formal) (PVF) resin as a scaffold. However, a low seeding efficiency of about 30% remains a major obstacle to the scaleup of the reactor. In the present study, a new cell seeding method, centrifugal cell immobilization (CCI), which is based on alternating centrifugation and resuspension, was used to achieve high-density seeding and improve the seeding efficiency. Using the CCI method, the maximum density of the immobilized hepatocytes reached 3.8 x 10(7) cells/1 cm(3)-PVF, and the seeding efficiency was improved to about 43% after a relatively short immobilization process (about 15 min). Moreover, further improvement of the seeding efficiency was obtained by serial immobilization procedures. Thus, we concluded that this method is useful and effective for seeding cells into 3-dimensional scaffolds.

VEGF Release in Multiluminal Hydrogels Directs Angiogenesis from Adult Vasculature In Vitro

“…6 Despite the angiogenic effect of such factors, comprehensive vascularization of thick tissues in vitro cannot be controlled and lack reproducibility. 23 Methods such as dynamic perfusion, 44,49 centrifugal force, 14,52 rotation, 31 magnetic cell seeding, 35 and stirring 22 have been used to achieve homogeneous cell distribution in 3-dimensional models of vasculogenesis. More recently, guided angiogenesis has been demonstrated using filamentous polymeric scaffolds.…”

Application of a polyelectrolyte complex coacervation method to improve seeding efficiency of bone marrow stromal cells in a 3D culture system

“…However, the efficiency and efficacy of dynamic seeding has been reported to be controversial [4,5,[7][8][9]. Various other seeding configurations have been developed to circumvent the limitations of static and dynamic seeding, which include filtration [3], oscillating perfusion [4], centrifuge [10] and perfusion cartridge seeding [11]. However, the application of these methods may be limited to scaffolds of a specific range of shape and morphology since most of the studies were performed on a single type of scaffold.…”

Seeding of mesenchymal stem cells into inner part of interconnected porous biodegradable scaffold by a new method with a filter paper

“…In addition, the flow seeding method has the advantage of preventing loss of inoculated cells outside of the scaffold in a 6-mm well or centrifuge tube. Compared to other methods 22,[31][32][33][34][35][36][37][38] , the flow method is simple, and can be carried out in usual Suspensions containing 1×10 6 cells were applied by the four methods. DNA was determined using a DNA kit (PicoGreen, Molecular Probe, Chicago, USA).…”